Therapy for complications of diabetes

ABSTRACT

A method for enhancing glycemic control and/or insulin sensitivity in a human subject having diabetic nephropathy and/or metabolic syndrome comprises administering to the subject a selective endothelin A (ET A ) receptor antagonist in a glycemic control and/or insulin sensitivity enhancing effective amount. A method for treating a complex of comorbidities in an elderly diabetic human subject comprises administering to the subject a selective ET A  receptor antagonist in combination or as adjunctive therapy with at least one additional agent that is (i) other than a selective ET A  receptor antagonist and (ii) effective in treatment of diabetes and/or at least one of said comorbidities other than hypertension. A therapeutic combination useful in such a method comprises a selective ET A  receptor antagonist and at least one antidiabetic, anti-obesity or antidyslipidemic agent other than a selective ET A  receptor antagonist.

CROSS-REFERENCE TO RELATED APPLICATIONS

This is a continuation from U.S. patent application Ser. No. 12/196,635,filed on Aug. 22, 2008, which claims priority to U.S. Provisional PatentApplication No. 60/957,300, filed on Aug. 22, 2007, the contents of eachof which are herein fully incorporated by reference.

This application claims the benefit of U.S. provisional application Ser.No. 60/957,300 filed on Aug. 22, 2007, the disclosure of which isincorporated herein by reference in its entirety. Further, thisapplication contains subject matter related to copending U.S.application Ser. No. 11/509,897, the disclosure of which is incorporatedherein by reference in its entirety without admission that suchdisclosure constitutes prior art to the present invention.

FIELD OF THE INVENTION

The present invention relates to methods and therapeutic combinationsuseful for improving clinical outcomes in diabetic patients havingcomplications of diabetes such as diabetic nephropathy and/or metabolicsyndrome.

BACKGROUND OF THE INVENTION

Hyperglycemia in diabetes mellitus, if not controlled, over time causescertain irreversible morphologic changes including glomerular fibrosisin kidneys of affected subjects, a condition known as diabeticnephropathy that is associated with decline in renal function,eventually leading to end-stage renal disease. In type 1(insulin-dependent) diabetes, glycemic control is usually achievablewith chronic insulin therapy; however, in type 2 (non-insulin-dependent)diabetes, insulin alone may be ineffective in preventing hyperglycemia.Even in patients with type 1 diabetes, insulin sensitivity can bepartially or completely lost. Insulin resistance, or loss of insulinsensitivity, is one of an array of physiological changes that occur insome individuals who are both obese and diabetic; such physiologicalchanges are collectively known as metabolic syndrome. Diabeticnephropathy and metabolic syndrome are serious complications of diabetesthat can dramatically reduce quality of life and survival time. Afeature of both these complications is arterial hypertension, whichsuperimposes risk of serious cardiac adverse events on the already highrisk of chronic kidney failure arising from these complications.

Endothelins (ETs), particularly ET-1, are believed to play a role inmediating the damaging effects of hyperglycemia in the kidney andelsewhere. Expression of ET-1 in endothelial cells of the renalvasculature is upregulated by hyperglycemia; the potent profibroticaction of ET-1 thus generated in the kidney is believed to be involvedin the morphologic changes seen in diabetic nephropathy. ET-1 acts viaendothelin A (ET_(A)) and endothelin B (ET_(B)) receptors. Elevatedplasma ET levels have been reported in patients with diabetes mellitus.See, for example, Takahashi et al. (1990) Diabetologia 33:306-310.

Elevated plasma ET levels have also been reported in patients withmetabolic syndrome. See Ferri et al. (1997) Exp. Clin. Endocrinol.Diabetes 105:38-40. Metabolic syndrome (sometimes referred to as“syndrome X”) is characterized by coexistence of glucose intolerance,hypertension, dyslipidemia (specifically elevated LDL (low densitylipoprotein) cholesterol and triglycerides and reduced HDL (high densitylipoprotein) cholesterol), obesity and susceptibility to cardiovasculardisease; these effects are thought to involve a common mechanism inwhich insulin resistance plays an important part.

The earliest clinical evidence of diabetic nephropathy ismicroalbuminuria, the appearance of low but abnormal levels (≧30 mg/day)of albumin in the urine. This early stage in development of the diseaseis known as incipient diabetic nephropathy. Without intervention, about80% of subjects with type 1 diabetes who develop sustainedmicroalbuminuria exhibit a progressive increase in urinary albumin,eventually (typically after 10-15 years) reaching clinical albuminuria(≧300 mg/day), a stage known as overt diabetic nephropathy. Accompanyingthe increase in albumin excretion is development of arterialhypertension. In subjects with overt diabetic nephropathy, withoutintervention, glomerular filtration rate (GFR) gradually falls over aperiod of 10-20 years, culminating in end-stage renal disease. SeeAmerican Diabetes Association (2004) Diabetes Care 27 (suppl.1):S79-S83. Structural changes in diabetic nephropathy include, in theincipient stage, mesangial expansion and a thickening of the glomerularbasement membrane (GBM). An increase in glomerular and kidney size isgenerally observed. Later, during the overt stage, mesangial nodules andtubular interstitial fibrosis develop.

Hocher et al. (2001) Nephron 87:161-169 reported that in rats withstreptozotocin-induced diabetes, administration of either the selectiveET_(A) receptor antagonist LU 135252 (darusentan) or the nonselectiveET_(A)/ET_(B) receptor antagonist LU 224332, in both cases at a dose of100 mg/kg/day, normalized glomerular matrix protein deposition,indicating an antifibrotic effect. However, neither compound was foundto influence serum glucose concentrations in the course of the study.

Dhien et al. (2000) J. Pharmacol. Exp. Therap. 293:351-359 reported thatLU 135252 at 100 mg/kg/day partially or fully reversed various renaleffects of streptozotocin-induced diabetes in rats, including increasedglomerular diameter and deposition of eosinophilic material within theglomeruli, but that plasma glucose levels were unaffected by LU 135252.

Sorokin & Kohan (2003) Am. J. Physiol. Renal Physiol. 285:F579-F589remarked that the stage was set for clinical trials of ET inhibitors inpatients with glomerular disease characterized by increased ET-1production and actions.

Avosentan, which may be classified as a selective ET_(A) or dualET_(A)/ET_(B) receptor antagonist, has been reported to be in Phase IIIclinical development for diabetic nephropathy. See Battistini et al.(2006) Exp. Biol. Med. 231:653-695.

U.S. Pat. No. 6,197,780 to Münter & Kirchengast reported that treatmentof obese mice with “substance 23” (darusentan) at 50 mg/kg/daycompletely prevented increase in body weight. A method is claimedtherein for treating a patient having hyperlipidemia, comprisingadministering a therapeutically effective amount of an ET antagonist(e.g., darusentan) to the patient.

Balsiger et al. (2002) Clin. Sci. 103 (Suppl. 48):4305-4335 reportedthat in a rat model of type 2 diabetes, BSF 208075 (said to be aselective ET_(A) receptor antagonist) reduced plasma glucose levels andimproved plasma glucose clearance rates in hyperglycemic rats.

On the other hand, Shaw et al. (2006) Exp. Biol. Med. 231:1101-1105reported that in a mouse model of non-obese type 1 diabetes, theselective ET_(A) receptor antagonist LU 208075 (ambrisentan) did notreduce the elevated plasma glucose levels seen in untreated animals.

According to Berthiaume et al. (2005) Metab. Clin. Exp. 54:735-740, somestudies have shown desensitization by ET-1 of insulin signaling, leadingto a decrease in glucose uptake, while other studies have shown oppositeresults. A study is reported therein of effects of the selective ET_(A)receptor antagonist atrasentan in a rat model of insulin resistance. Ata dose of 5 mg/kg/day, atrasentan was reported to significantly reduce3-hour fasting insulin level but not 3-hour fasting glucose level, andto significantly reduce Δ_(AUC), a measure of incremental area under thecurve induced by a meal tolerance test, for glucose, insulin andglucose-insulin index. These results were said to demonstrate animprovement in glucose tolerance and insulin sensitivity and to suggestthat chronic endothelin antagonism may have benefits in treatment ofinsulin resistance and/or diabetes. It was further reported that ET_(A)receptor blockade by atrasentan led to an increase rather than adecrease in plasma ET-1 levels.

Shaw & Boden (2005) Current Vascular Pharmacology 3:359-363 reviewedevidence on effects of ET-1 and proposed that chronically elevated ET-1levels may be a cause of insulin resistance and impaired glucosetolerance in early stages of type 2 diabetes, obesity and metabolicsyndrome. Recent data were said therein to indicate that combinedET_(A)/ET_(B) receptor antagonists may function as effectively asselective ET_(A) blockers. A need was proposed for prospective trials toassess whether ET-1 antagonists, either alone or in combination, aresuperior to other more conventional treatments such as insulinsensitizers and to evaluate effects of combined therapies on developmentof insulin resistance and progression of diabetes.

Subjects having diabetic nephropathy and/or metabolic syndrome representa particularly challenging subpopulation of diabetic patients, for whomtherapies giving improved outcomes with respect to quality of life andsurvival time, through enhanced glycemic control and/or insulinsensitivity, would represent an important advance in the art.

Recognizing that elevated blood pressure occurs in both diabeticnephropathy and metabolic syndrome, and brings its own attendant risksto quality of life and survival time, an even more challenging patientpopulation comprises subjects having at least one of these complicationsof diabetes and exhibiting inadequate blood pressure control by standardantihypertensive therapies. Subjects exhibiting resistance to a baselineantihypertensive therapy with one or more drugs include patients havingclinically diagnosed resistant hypertension. Resistant hypertension isdefined by the Seventh Report of the Joint National Committee onPrevention, Detection, Evaluation, and Treatment of High Blood Pressure(JNC 7; Chobanian et al. (2003) Hypertension 42:1206-1252) as a failureto achieve goal blood pressure in patients who are adhering to fulldoses of an appropriate three-drug regimen that includes a diuretic.Further, resistant hypertension is diagnosed by many physicians on thebasis of a patient's resistance to adequate, but less than full, dosesof an appropriate three-drug regimen because of the risk or occurrenceof adverse events associated with full doses. The terms “adequate” and“full” in the present context are defined hereinbelow.

For patients with serious or compelling conditions such as diabetes andchronic kidney disease, JNC 7 recommends a goal of systolic bloodpressure (SBP)<130 mmHg and diastolic blood pressure (DBP)<80 mmHg.Despite intensive, multi-drug therapy, only about 50% of patients withdiabetes or chronic kidney disease reach traditional blood pressuregoals, with even fewer reaching the more stringent goals now recommendedby JNC 7. Thus, resistant hypertension is particularly acute forsegments of the population which exhibit complications of diabetes suchas diabetic nephropathy or metabolic syndrome.

It should be noted that the British Hypertensive Society (BHD-IV; J.Human Hypertens. (2004) 18:139-185), the European Society ofHypertension/European Society of Cardiology (ESH/ESC; J. Hypertens.(2003) 21:1011-1053), and the World Health Organization/InternationalSociety of Hypertension (WHO/ISH; J. Hypertens. (2003) 21:1983-1992)guidelines propose similar but not identical blood pressure goals fordiabetic patients.

In a news release dated Aug. 18, 2005(http://stockjunction.com/modules.php?name=News&file=print&sid=7332),Myogen Inc. reported positive results in a clinical trial (DAR-201)evaluating darusentan in resistant hypertension. Among inclusioncriteria for DAR-201 were subjects with diabetes and/or chronic kidneydisease with mean systolic blood pressure ≧130 mmHg(http://clinicaltrials.gov/ct/show/NCT00364026).

Weber et al. (2006) presented a poster, available athttp://www.secinfo.com/dvjdn.vbz.d.htm, posted May 16, 2006, reporting,inter alfa, subject demographics in the DAR-201 study. Of 115 subjectsenrolled, 70 had diabetes and/or chronic kidney disease, 55 had diabetesand 29 had chronic kidney disease.

Nakov et al. (2002) Am. J. Hypertens. 15:583-589 described a 392-patientstudy in which moderate hypertension was treated with darusentan at 10to 100 mg/day. Exclusion criteria included concomitant medication withother antihypertensive drugs. Darusentan was reported to significantlyreduce SBP and DBP by comparison with placebo.

German Patent No. DE 19744799 of Knoll mentions, in the abstractthereof, combinations of an endothelin antagonist, such as darusentan,and a diuretic said to show synergistic activity in treatment ofhypertension, coronary artery disease, cardiac or renal insufficiency,renal or myocardial ischemia, subarachnoid hemorrhage, Raynaud's diseaseand peripheral arterial occlusion.

U.S. Pat. No. 6,352,992 to Kirchengast et al. proposes pharmaceuticalcombination preparations comprising a beta-receptor blocker and anendothelin antagonist for treatment of vasoconstrictive disorders. Amongendothelin antagonists mentioned is darusentan.

German Patent No. DE 19743142 of Knoll proposes combinations of anendothelin antagonist, such as darusentan, and a calcium antagonist fortreatment of cardiovascular disorders such as pulmonary hypertension andrenal and myocardial ischemia.

U.S. Pat. No. 6,329,384 to Munter et al. proposes combinations ofendothelin antagonists, such as darusentan, and renin-angiotensin systeminhibitors, in particular angiotensin II antagonists and angiotensinconverting enzyme (ACE) inhibitors for treatment of vasoconstrictivedisorders such as hypertension, heart failure, ischemia or vasospasms.

German Patent No. DE 19743140 of Knoll proposes combinations of anendothelin antagonist, such as darusentan, and a vasodilator fortreatment of cardiovascular disorders such as pulmonary hypertension,renal or myocardial ischemia, subarachnoid hemorrhage, Raynaud'sdisease, and peripheral arterial occlusion.

International Patent Publication No. WO 2004/082637 of Pharmaciaproposes combinations of an aldosterone receptor antagonist with anendothelin receptor antagonist and/or an endothelin converting enzymeinhibitor, compositions thereof, and therapeutic methods for use intreatment of pathological conditions such as hypertension,cardiovascular disease and renal dysfunction.

Improved drug therapies for treatment of patients having complicationsof diabetes such as diabetic nephropathy and/or metabolic syndrome,especially such patients exhibiting resistance to a baselineantihypertensive therapy with one or more drugs, for example patientshaving clinically diagnosed resistant hypertension, would be highlydesirable.

SUMMARY OF THE INVENTION

There is now provided a method for enhancing glycemic control and/orinsulin sensitivity in a human subject having diabetic nephropathyand/or metabolic syndrome, comprising administering to the subject aselective endothelin A (ET_(A)) receptor antagonist in a glycemiccontrol and/or insulin sensitivity enhancing effective amount.

There is further provided a method for treating a complex ofcomorbidities in an elderly diabetic human subject, comprisingadministering to the subject a selective ET_(A) receptor antagonist incombination or adjunctive therapy with at least one additional agentthat is (i) other than a selective ET_(A) receptor antagonist and (ii)effective in treatment of diabetes and/or at least one of saidcomorbidities other than hypertension. Optionally the combination oradjunctive therapy further comprises administration of at least oneantihypertensive other than a selective ET_(A) receptor antagonist.

There is still further provided a therapeutic combination comprising aselective ET_(A) receptor antagonist and at least one antidiabetic,anti-obesity or antidyslipidemic agent other than a selective ET_(A)receptor antagonist.

Other embodiments, including particular aspects of the embodimentssummarized above, will be evident from the detailed description thatfollows.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic diagram of the clinical study described in Example2 herein.

DETAILED DESCRIPTION

The term “diabetic nephropathy” as used herein will be understood toinclude both incipient and overt stages of diabetic nephropathy, whetherdiagnosed or not, but most typically as diagnosed by a clinician orphysician. The term “metabolic syndrome” as used herein refers to acomplex of obesity, hypertension, dyslipidemia and diabetes marked by adegree of insulin resistance. The existence of metabolic syndrome as atrue clinical syndrome is not universally accepted; it will beunderstood that in the present context a patient having metabolicsyndrome is one exhibiting a complex of conditions as itemized above,whether or not “metabolic syndrome” is formally diagnosed in thepatient.

In one embodiment, the method of the invention is “for enhancingglycemic control and/or insulin sensitivity” in a human subject. Whereglycemic control is enhanced, such enhancement can, but does notnecessarily, arise from increased insulin sensitivity. Likewise, whereinsulin sensitivity is enhanced, such enhancement can, but does notnecessarily, lead to improved glycemic control.

In one embodiment, practice of the method leads to enhancement ofglycemic control, particularly in a subject having diabetic nephropathyand/or metabolic syndrome.

In another embodiment, practice of the method leads to enhancement ofinsulin sensitivity, particularly in a subject having diabeticnephropathy and/or metabolic syndrome. Insulin sensitivity may bemeasured by standard insulin models, such as HOMA-IR (homeostasis modelassessment of insulin resistance).

In yet another embodiment, practice of the method leads to enhancementin both glycemic control and insulin sensitivity, particularly in asubject having diabetic nephropathy and/or metabolic syndrome.

Enhancement of glycemic control is typically manifested by a reductionin tendency for hyperglycemia. A fasting (e.g., preprandial) bloodglucose level greater than about 140 mg/dl, or a bedtime blood glucoselevel greater than about 160 mg/dl, can be evidence of hyperglycemia.Any reduction in blood glucose level can be beneficial to a subjecthaving hyperglycemia, for example a reduction by at least about 5, atleast about 10, at least about 15 or at least about 20 mg/dl. Ideally,glucose level is brought into a goal range for healthy subjects of about80 to about 120 mg/dl (preprandial) or about 100 to about 140 mg/dl(bedtime). Reduction of glucose level in urine can also provide evidenceof enhanced glycemic control, but is less reliable than bloodmeasurements because excretion of glucose in urine typically does notoccur unless blood glucose level exceeds about 180 mg/dl.

For some purposes, a superior measure of glycemic control is theglycosylated hemoglobin (HbA_(1c)) test. This test reflects bloodglucose concentration over a period of time related to the life-span ofred blood cells (about 120 days), and consequently is not affected bydaily or hourly fluctuations in blood glucose level. A patient having anHbA_(1c) test result greater than about 8% is normally consideredhyperglycemic. Any reduction in HbA_(1c) level can be beneficial to sucha patient, for example a reduction by at least about 0.5, at least about1, at least about 1.5 or at least about 2 percentage points. Ideally,HbA_(1c) level is brought into a goal range for healthy subjects ofabout 4% to about 6%.

Subjects having diabetic nephropathy and/or metabolic syndrome can be ofany age, but incidence of these complications of diabetes increasesmarkedly with age. Older subjects can respond differently from youngersubjects to treatment, and can have a different spectrum of adverseeffects. It is contemplated herein that elderly subjects (i.e., subjectsat least about 50, for example at least about 55, at least about 60 orat least about 65 years old) can benefit especially greatly fromtreatment according to the present method, in part because of theseverity with which their quality of life and survival time are impactedby these complications of diabetes, and in part because the particularadverse side effects that have been noted for ET antagonists, includingreproductive effects, are of lesser significance later in life.

In one embodiment the subject has incipient diabetic nephropathy.

In another embodiment the subject has overt diabetic nephropathy.

In a further embodiment the subject has incipient or overt diabeticnephropathy and practice of the method provides a further beneficialeffect in one or more morphologic markers of diabetic nephropathy. A“morphologic marker” in the present context is any structural orhistological feature of the kidney or a tissue thereof, whetherobservable macroscopically, by light microscopy or by electronmicroscopy. Such markers can be observable directly, for example bybiopsy, or indirectly, through a secondary effect specific to themarker. Examples of morphologic markers of diabetic nephropathy include,without limitation, kidney size, kidney weight, GBM thickening,mesangial expansion, deposition of collagen, fibronectin and laminin,nephron density, nodular glomerulosclerosis, atherosclerosis of renalvasculature or a combination thereof.

In a still further embodiment the subject has incipient or overtdiabetic nephropathy and practice of the method provides a furtherbeneficial effect in one or more indicators of renal function, forexample GFR, creatinine clearance, albuminuria or a combination thereof.

A “beneficial effect” of the present method can take the form of animprovement over baseline, i.e., an improvement over a measurement orobservation made prior to initiation of therapy according to the method.For example, a reduction in amount of collagen deposited, an increase inGFR (in a subject having overt diabetic nephropathy) or a reduction inalbuminuria by comparison with a baseline level would represent such animprovement.

A “beneficial effect” can also take the form of an arresting, slowing,retarding or stabilizing of a deleterious progression in any of theabove markers or functional effects of diabetic nephropathy. Forexample, even if GFR is not increased or albuminuria is not reduced bycomparison with baseline measurements, a reduction in the rate ofdecrease of GFR or the rate of increase of albuminuria would represent abeneficial effect of the present method.

Thus in one embodiment, practice of the invention leads to an arresting,slowing, retarding or stabilizing of the progression of diabeticnephropathy in the subject. Such arresting, slowing, retarding orstabilizing of progression of the disease can result in extension of thetime to end-stage renal disease or chronic kidney failure, which in turncan extend survival time of the subject.

A selective ET_(A) receptor antagonist useful herein exhibits anaffinity (as expressed by dissociation constant K_(i)) for ET_(A) notgreater than about 10 nM and a selectivity for ET_(A) over ET_(B) (asexpressed by the ratio K_(i)(ET_(B))/K_(i)(ET_(A))) of at least about50. In various embodiments K_(i)(ET_(A)) is not greater than about 5 nM,not greater than about 2 nM, not greater than about 1 nM, not greaterthan about 0.5 nM or not greater than about 0.2 nM. In variousembodiments K_(i)(ET_(B))/K_(i)(ET_(A)) is at least about 100, at leastabout 200, at least about 500 or at least about 1000.

Suitable selective ET_(A) receptor antagonists can be identified by oneof ordinary skill from literature on such antagonists, based on thedisclosure herein, but a non-limiting list of such antagonists includesambrisentan, atrasentan, avosentan, BMS 193884, BQ-123, CI-1020,clazosentan, darusentan, edonentan, S-0139, SB-209670, sitaxsentan,TA-0201, tarasentan, TBC 3711, tezosentan, YM-598, ZD-1611 and ZD-4054,as well as salts, esters, prodrugs, metabolites, tautomers, racematesand enantiomers thereof.

In one embodiment, the selective ET_(A) receptor antagonist comprisesdarusentan((+)-(S)-2-[(4,6-dimethoxy-2-pyrimidinyl)oxy]-3-methoxy-3,3-diphenylpropionicacid). This compound has the formula

Riechers et al. (1996) J. Med. Chem. 39:2123-2128 reported that2-(4,6-dimethoxypyrimidin-2-yloxy)-3-methoxy-3,3-diphenylpropionic acidin racemic form has K_(i)(ET_(A)) of 6 nM and K_(i)(ET_(B)) of 371 nM,thus selectivity K_(i)(ET_(B))/K_(i)(ET_(A)) for the racemate based onthese data can be calculated as about 62. It was further reportedtherein that the pure enantiomers have an affinity for ET_(A) of 3 nMand 150 nM. The more potent enantiomer was concluded to be the(S)-enantiomer.

More recently, Lip (2001) IDrugs 4(11):1284-1292 reported the(S)-enantiomer (darusentan) as having K_(i)(ET_(A)) 1.4 nM andselectivity K_(i)(ET_(B))/K_(i)(ET_(A)) about 160.

It has now been found that K_(i)(ET_(B))/K_(i)(ET_(A)) for darusentan,when measured in a system that achieves steady-state binding, is muchgreater than previously reported.

To measure affinities of darusentan for ET_(A) and ET_(B) receptors inthe same human tissue preparation, ¹²⁵[I]-endothelin-1 receptor bindingcold ligand competition curves were performed in human myocardialmembranes prepared from failing and non-failing left ventricles, andcold ligand dissociation constants (K_(i)) for ET_(A) and ET_(B)receptors were determined by computer modeling. Assay conditionsincluded 10 μM Gpp(NH)p (guanylyl-5′-imidodiphosphate) to eliminatehigh-affinity agonist binding, 18-point competition curves from 1 pM to100 μM, and a 4-hour incubation time to achieve steady-state binding.Darusentan was found to have the following properties under theseconditions (mean of 8 assays):

-   -   K_(i)(ET_(A)): 0.178±0.055 nM    -   K_(i)(ET_(B)): 216±85 nM    -   K_(i)(ET_(B))/K_(i)(ET_(A)): 1181±148

According to the present method, the selective ET_(A) receptorantagonist is administered “in a glycemic control and/or insulinsensitivity enhancing effective amount.” What constitutes such aneffective amount will depend on the particular selective ET_(A) receptorantagonist used, on the individual subject, on the dosage form and routeof administration used and on other factors, and can be readilydetermined by one of skill in the art without undue experimentationbased on the disclosure herein. Any dose that is effective for enhancingglycemic control and/or insulin sensitivity, up to a maximum that istolerated by the subject without unacceptable adverse side effects, canbe administered. Illustratively for darusentan, such a dose is likely tobe about 1 to about 600 mg/day, for example about 5 to about 450 mg/dayor about 10 to about 300 mg/day. Higher or lower doses can be useful inspecific circumstances. Useful doses of other selective ET_(A) receptorantagonists are doses that are therapeutically equivalent to such a doseof darusentan.

The desired daily dosage amount can be administered in any suitablenumber of individual doses, for example four times, three times, twiceor once a day. With a dosage form having appropriate controlled releaseproperties, a lower frequency of administration may be possible, forexample once every two days, once a week, etc.

In one embodiment, the selective ET_(A) receptor antagonist isadministered according to a therapeutic regimen wherein dose andfrequency of administration and duration of therapy are effective tolower blood glucose level by at least about 10 mg/dl and/or to lowerHbA_(1c) level by at least about 0.5 percentage points.

In another embodiment, the selective ET_(A) receptor antagonist isadministered according to a therapeutic regimen wherein dose andfrequency of administration and duration of therapy are effective toachieve a goal preprandial blood glucose level of about 80 to about 120mg/dl, a goal bedtime blood glucose level of about 100 to about 140mg/dl and/or a goal HbA_(1c) level not greater than about 7%.

Benefits of the present method may not be evident immediately uponinitiating a therapeutic regimen as described herein. In particular, itcan take some time for it to become evident that progression of acomplication of diabetes such as diabetic neuropathy has been slowed. Itis therefore contemplated that administration of a selective ET_(A)receptor antagonist will in some cases continue for an extended periodof time, typically at least about 1 month, more typically at least about3 months. Duration of therapy in some embodiments can be at least about1 year, at least about 5 years, or for as long as needed, which can belifelong (i.e., from a time of initiation of treatment for substantiallythe remainder of the patient's life). In one embodiment duration oftherapy is from a time of diagnosis of diabetic nephropathy (whetherincipient or overt) at least to a time of progression of the diabeticnephropathy into end-stage renal disease. In some situations the methodof this embodiment will be successful in preventing progression of thedisease to end-stage; in such situations the treatment can be continuedindefinitely. For example, administration of the selective ET_(A)receptor antagonist can continue for as long as a therapeutic benefit isprovided thereby and any adverse side effects thereof remaincommensurate with the therapeutic benefit.

Selective ET_(A) receptor antagonists are known to be useful asantihypertensive agents. Thus practice of the present method is likelyto provide a benefit not only in glycemic control and/or insulinsensitivity as outlined above, but an additional benefit in loweringblood pressure. As hypertension is an important feature of both diabeticnephropathy and metabolic syndrome, these benefits can be mutuallyreinforcing.

Any one or more measures of blood pressure can be lowered by a method asdescribed herein, including SBP and/or DBP as determined, for example,by sphygmomanometry. SBP and/or DBP can be measured, for example, in asitting or ambulatory patient.

A “trough sitting” SBP or DBP is measured at a time point when serumconcentration of a drug or drugs administered according to a method ofthe invention is expected to be at or close to its lowest in a treatmentcycle, typically just before administration of a further dose.Illustratively, where the drug or drugs are administered once a day at aparticular time, for example around 8 am, trough sitting systolic ordiastolic blood pressure can be measured at that time, immediatelybefore the daily administration. It is generally preferred to measuretrough sitting SBP or DBP at around the same time of day for each suchmeasurement, to minimize variation due to the natural diurnal bloodpressure cycle.

A “24-hour ambulatory” SBP or DBP is an average of measurements takenrepeatedly in the course of a 24-hour period, in an ambulatory patient.

A “maximum diurnal” SBP or DBP is a measure of highest SBP or DBPrecorded in a 24-hour period, and often reflects the peak of the naturaldiurnal blood pressure cycle, typically occurring in the morning, forexample between about 5 am and about 11 am. Commonly, a second peakoccurs in the evening, for example between about 5 pm and 10 pm. Such abimodal waveform diurnal ABP pattern may be especially characteristic ofresistant hypertension.

A common feature of resistant hypertension is a nighttime (definedherein as 2200 to 0600) mean systolic ABP that is less than about 10%lower than the daytime (defined herein as 0600 to 2200) mean systolicABP. The parameter herein termed “day/night ABP ratio” expressed as apercentage is calculated as (daytime mean−nighttime mean)/daytimemean×100. A diurnal ABP pattern having a day/night ABP ratio of lessthan about 10% is sometimes referred to as a “non-dipping ABP”.

The patient receiving therapy according to a method of the invention canbe a patient exhibiting resistance to a baseline antihypertensivetherapy with one or more drugs. A “baseline antihypertensive therapy”herein means a therapeutic regimen comprising administration of one ormore drugs, not including a selective ET_(A) receptor antagonist, withan objective (which can be the primary objective or a secondaryobjective of the regimen) of lowering blood pressure in the patient.Each drug according to the regimen is administered at least at a doseconsidered by an attending physician to be adequate for treatment ofhypertension, taking into account the particular patient's medicalcondition and tolerance for the drug without unacceptable adverseside-effects. An “adequate” dose as prescribed by the physician can beless than or equal to a full dose of the drug. A “full” dose is thelowest of (a) the highest dose of the drug labeled for a hypertensionindication; (b) the highest usual dose of the drug prescribed accordingto JNC 7, BHD-IV, ESH/ESC or WHO/ISH guidelines; or (c) the highesttolerated dose of the drug in the particular patient.

A baseline antihypertensive therapy illustratively comprisesadministering one or more diuretics and/or one or more antihypertensivedrugs selected from (a) angiotensin converting enzyme inhibitors andangiotensin II receptor blockers, (b) beta-adrenergic receptor blockers,(c) calcium channel blockers, (d) direct vasodilators, (e)alpha-1-adrenergic receptor blockers, (f) central alpha-2-adrenergicreceptor agonists and other centrally acting antihypertensive drugs, and(g) aldosterone receptor antagonists. Optionally drugs of still furtherclasses can be included in the baseline therapy.

A patient who is “resistant” to a baseline antihypertensive therapy isone in whom hypertension is failing to respond adequately or at all tothe baseline therapy. Typically, the patient receiving the baselinetherapy is failing to reach an established blood pressure goal, as setforth for U.S. patients, for example, in JNC 7 or comparable standardsin other countries (e.g., BHD-IV, ESH/ESC or WHO/ISH guidelines).Illustratively, the JNC 7 goal in a patient having a complicatingcondition such as diabetes and/or chronic kidney disease is <130 mmHgSBP and <80 mmHg DBP.

Patients resistant to a baseline antihypertensive therapy, especiallysuch a therapy involving a plurality of drugs, clearly represent a verychallenging population for treatment. Typically in such patients,increasing dosages of the baseline therapy are not an option because ofresulting adverse side effects; furthermore this approach is oftenineffective in providing a desired lowering of blood pressure.

A clinically meaningful lowering of blood pressure can be obtained insuch patients by use of a selective ET_(A) receptor antagonist such asdarusentan. A reduction of at least about 3 mmHg in any blood pressureparameter can be considered clinically meaningful.

Accordingly, in one embodiment of the present invention, a method forenhancing glycemic control and/or insulin sensitivity and for loweringblood pressure in a patient exhibiting resistance to a baselineantihypertensive therapy comprises administering to the patient aselective ET_(A) receptor antagonist, for example darusentan, at a doseand frequency effective to provide a reduction of at least about 3 mmHgin trough sitting SBP and/or DBP, 24-hour ambulatory SBP and/or DBP,and/or maximum diurnal SBP and/or DBP.

In patients exhibiting resistance to a baseline antihypertensive therapywith one or more drugs, administration of darusentan adjunctively withthese same drugs is surprisingly well tolerated. Accordingly, in anotherembodiment of the present invention, a method for enhancing glycemiccontrol and/or insulin sensitivity and for lowering blood pressure in apatient exhibiting resistance to a baseline antihypertensive therapywith one or more drugs comprises administering darusentan to the patientadjunctively with said one or more drugs.

While in certain embodiments the selective ET_(A) receptor antagonist,for example darusentan, can be administered alone, i.e., in monotherapy,it is contemplated that in most cases combination therapy, for examplebut not necessarily with one or more drugs of the baseline therapy towhich the patient has proved resistant, will be desirable. However, abenefit of the administration of darusentan can be that, at least insome circumstances, it can permit dose reduction, or even elimination,of at least one of the drugs in the baseline therapy.

Particularly when used at a full dose, many baseline antihypertensivetherapy drugs can have undesirable, in some cases clinicallyunacceptable or even dangerous, adverse side effects.

For example, especially at full doses, potassium-sparing diuretic drugscan be associated with increased risk of hyperkalemia and relateddisorders. Overuse of loop diuretics can cause depletion of sodiumresulting in hyponatremia and/or extracellular fluid volume depletionassociated with hypotension, reduced GFR, circulatory collapse, andthromboembolic episodes. Further, loop diuretics can cause ototoxicitythat results in tinnitus, hearing impairment, deafness and/or vertigo.Thiazide diuretics, similarly to loop diuretics, can have adverseeffects related to abnormalities of fluid and electrolyte balance. Suchadverse events include extracellular volume depletion, hypotension,hypokalemia, hyponatremia, hypochloremia, metabolic alkalosis,hypomagnesemia, hypercalcemia and hyperuricemia. Thiazide diuretics canalso decrease glucose tolerance, and increase plasma levels of LDLcholesterol, total cholesterol, and total triglycerides.

Angiotensin converting enzyme (ACE) inhibitors are associated with coughand increased risk of angioedema. Beta-adrenergic receptor blockers areassociated with increased risk of bronchospasm, bradycardia, heartblock, excess negative inotropic effect, peripheral arterialinsufficiency and sometimes male impotence. Calcium channel blockers areassociated with increased risk of lower limb edema. Further informationon adverse events associated with antihypertensive drugs can be found,for example, in standard reference works such as Goodman & Gilman's ThePharmaceutical Basis of Therapeutics, 13th ed.

In situations such as those outlined immediately above, dose reductionor elimination of a baseline therapy drug permitted by use of darusentancan result in a reduced risk or incidence of adverse events bycomparison with the baseline therapy alone without such dose reductionor elimination.

“Adjunctive” administration of darusentan (or other selective ET_(A)receptor antagonist) in the present context means that the darusentan isadministered concomitantly with a baseline hypertensive therapy asdefined above, with or without dose reduction of one or more drugs inthe baseline therapy. For example, darusentan can be administeredadjunctively with an adequate to full dose of each of the drugs in thebaseline therapy. In adjunctive therapy, the dose and frequency ofdarusentan administration is, in one embodiment, effective incombination with the baseline therapy to provide a reduction of at leastabout 3 mmHg in trough sitting SBP and/or DBP, 24-hour ambulatory SBPand/or DBP, and/or maximum diurnal SBP and/or DBP.

A method of the present invention is especially beneficial where thepatient has clinically diagnosed resistant hypertension. By definitionherein, in general accordance with JNC 7, such a patient exhibitsresistance to an antihypertensive regimen of at least three drugsincluding a diuretic. In one embodiment, the patient having resistanthypertension exhibits resistance to a baseline antihypertensive therapythat comprises at least the following:

-   -   (1) one or more diuretics; and    -   (2) two or more antihypertensive drugs, selected from at least        two of the following classes:        -   (a) ACE inhibitors and angiotensin II receptor blockers;        -   (b) beta-adrenergic receptor blockers; and        -   (c) calcium channel blockers.

In some cases, the patient is resistant to an even more comprehensivebaseline therapy, further comprising, for example, one or more directvasodilators, alpha-1-adrenergic blockers, central alpha-2-adrenergicagonists or other centrally acting antihypertensive drugs, and/oraldosterone receptor antagonists.

While in one embodiment the selective ET_(A) receptor antagonist isadministered orally, the invention is not limited to any route ofadministration, so long as the route selected results in effectivedelivery of the drug so that the stated benefits are obtainable. Thusadministration of the darusentan can illustratively be parenteral (e.g.,intravenous, intraperitoneal, subcutaneous or intradermal), transdermal,transmucosal (e.g., buccal, sublingual or intranasal), intraocular,intrapulmonary (e.g., by inhalation) or rectal. Most conveniently forthe majority of patients, however, the selective ET_(A) receptorantagonist is administered orally, i.e., per os (p.o.). Any suitableorally deliverable dosage form can be used for the selective ET_(A)receptor antagonist, including without limitation tablets, capsules(solid- or liquid-filled), powders, granules, syrups and other liquids,etc.

Most antihypertensive medicines are suitable for once a dayadministration, and this is true also of darusentan. Thus, particularlywhere darusentan is being administered in adjunctive therapy with one ormore other antihypertensive drugs, it is generally most convenient toadminister the darusentan once a day in a dose as indicated above.

Most typically, where the patient has clinically diagnosed resistanthypertension, the selective ET_(A) receptor antagonist is administeredadjunctively with (1) one or more diuretics; and (2) two or moreantihypertensive drugs, selected from (a) ACE inhibitors and angiotensinII receptor blockers; (b) beta-adrenergic receptor blockers; and (c)calcium channel blockers. Each of these diuretic and antihypertensivedrugs is typically administered at an adequate to full dose. One ofskill in the art can readily identify a suitable dose for any particulardiuretic or antihypertensive drug from publicly available information inprinted or electronic form, for example on the internet.

Mention of a particular diuretic or antihypertensive drug in the presentspecification and claims will be understood, except where the contextdemands otherwise, to include pharmaceutically acceptable salts, esters,prodrugs, metabolites, racemates and enantiomers of the drug, to theextent that such salts, esters, prodrugs, metabolites, racemates orenantiomers exist and are therapeutically effective.

Examples of drugs useful in combination or adjunctive therapy with aselective ET_(A) receptor antagonist, for example darusentan, or as acomponent of a baseline antihypertensive therapy are classified andpresented in several lists below. Some drugs are active at more than onetarget; accordingly certain drugs may appear in more than one list. Useof any listed drug in a combination or adjunctive therapy of theinvention is contemplated herein, independently of its mode of action.

A suitable diuretic can illustratively be selected from the followinglist:

Organomercurials

chlormerodrin

chlorothiazide

chlorthalidone

meralluride

mercaptomerin sodium

mercumatilin sodium

mercurous chloride

mersalyl

Purines

pamabrom

protheobromine

theobromine

Steroids

canrenone

oleandrin

spironolactone

Sulfonamide Derivatives

acetazolamide

ambuside

azosemide

bumetanide

butazolamide

chloraminophenamide

clofenamide

clopamide

clorexolone

disulfamide

ethoxzolamide

furosemide

mefruside

methazolamide

piretanide

torsemide

tripamide

xipamide

Thiazides and Analogs

althiazide

bendroflumethiazide

benzthiazide

benzylhydrochlorothiazide

buthiazide

chlorthalidone

cyclopenthiazide

cyclothiazide

ethiazide

fenquizone

hydrochlorothiazide

hydroflumethiazide

indapamide

methylclothiazide

metolazone

paraflutizide

polythiazide

quinethazone

teclothiazide

trichloromethiazide

Uracils

aminometradine

Unclassified

amiloride

Biogen BG 9719

chlorazanil

ethacrynic acid

etozolin

isosorbide

Kiowa Hakko KW 3902

mannitol

muzolimine

perhexyline

Sanofi-Aventis SR 121463

ticrynafen

triamterene

urea

In some embodiments, the diuretic if present comprises a thiazide orloop diuretic. Thiazide diuretics are generally not preferred where thepatient has a complicating condition such as diabetes or chronic kidneydisease, and in such situations a loop diuretic can be a better choice.

Particularly suitable thiazide diuretics, for example for use withdarusentan, include chlorothiazide, chlorthalidone, hydrochlorothiazide,indapamide, metolazone, polythiazide and combinations thereof.Particularly suitable loop diuretics include bumetanide, furosemide,torsemide and combinations thereof.

A suitable ACE inhibitor can illustratively be selected from thefollowing list:

alacepril

benazepril

captopril

ceronapril

cilazapril

delapril

enalapril

enalaprilat

eosinopril

fosinopril

imidapril

lisinopril

moexipril

moveltipril

omapatrilat

perindopril

quinapril

ramipril

sampatrilat

spirapril

temocapril

trandolapril

Particularly suitable ACE inhibitors, for example for use withdarusentan, include benazepril, captopril, enalapril, fosinopril,lisinopril, moexipril, perindopril, quinapril, ramipril, trandolapriland combinations thereof.

A suitable angiotensin II receptor blocker can illustratively beselected from the following list:

candesartan

eprosartan

irbesartan

losartan

olmesartan

tasosartan

telmisartan

valsartan

A suitable beta-adrenergic receptor blocker can illustratively beselected from the following list:

AC 623

acebutolol

alprenolol

atenolol

amosulalol

arotinolol

atenolol

befunolol

betaxolol

bevantolol

bisoprolol

bopindolol

bucindolol

bucumolol

bufetolol

bufuralol

bunitrolol

bupranolol

butidrine hydrochloride

butofilolol

carazolol

carteolol

carvedilol

celiprolol

cetamolol

cloranolol

dilevalol

esmolol

indenolol

labetalol

landiolol

levobunolol

mepindolol

metipranolol

metoprolol

moprolol

nadolol

nadoxolol

nebivolol

nifenalol

nipradilol

oxprenolol

penbutolol

pindolol

practolol

pronethalol

propranolol

sotalol

sulfinalol

talinolol

tertatolol

tilisolol

timolol

toliprolol

xibenolol

Particularly suitable beta-adrenergic receptor blockers, for example foruse with darusentan, include acebutolol, atenolol, betaxolol,bisoprolol, carvedilol, labetalol, metoprolol, nadolol, penbutolol,pindolol, propranolol, timolol and combinations thereof.

A suitable calcium channel blocker can illustratively be selected fromthe following list:

Arylalkylamines

bepridil

clentiazem

diltiazem

fendiline

gallopamil

mibefradil

prenylamine

semotiadil

terodiline

verapamil

Dihydropyridine Derivatives

amlodipine

aranidipine

barnidipine

benidipine

cilnidipine

efonidipine

elgodipine

felodipine

isradipine

lacidipine

lercanidipine

manidipine

nicardipine

nifedipine

nilvadipine

nimodipine

nisoldipine

nitrendipine

NZ 105

Piperazine Derivatives

cinnarizine

dotarizine

flunarizine

lidoflazine

lomerizine

Unclassified

bencyclane

etafenone

fantofarone

monatepil

perhexyline

Particularly suitable calcium channel blockers, for example for use withdarusentan, include amlodipine, diltiazem, felodipine, isradipine,nicardipine, nifedipine, nisoldipine, verapamil and combinationsthereof.

Optionally, one or more additional antihypertensive drugs can beadministered. These can be selected, for example, from directvasodilators, alpha-1-adrenergic receptor blockers, centralalpha-2-adrenergic receptor agonists and other centrally actingantihypertensive drugs, and aldosterone receptor antagonists.

A suitable direct vasodilator can illustratively be selected from thefollowing list:

amotriphene

benfurodil hemisuccinate

benziodarone

chloracizine

chromonar

clobenfurol

clonitrate

cloricromen

dilazep

droprenilamine

efloxate

erythrityl tetranitrate

etafenone

fendiline

hexestrol bis(β-diethylaminoethyl ether)

hexobendine

hydralazine

isosorbide dinitrate

isosorbide mononitrate

itramin tosylate

khellin

lidoflazine

mannitol hexanitrate

minoxidil

nitroglycerin

pentaerythritol tetranitrate

pentrinitrol

perhexyline

pimethylline

prenylamine

propatyl nitrate

trapidil

tricromyl

trimetazidine

trolnitrate phosphate

visnadine

Particularly suitable direct vasodilators, for example for use withdarusentan, include hydralazine, minoxidil and combinations thereof.

A suitable alpha-1-adrenergic receptor blocker can illustratively beselected from the following list:

amosulalol

arotinolol

carvedilol

dapiprazole

doxazosin

ergoloid mesylates

fenspiride

idazoxan

indoramin

labetalol

methyldopa

monatepil

naftopidil

nicergoline

prazosin

tamsulosin

terazosin

tolazoline

trimazosin

yohimbine

Particularly suitable alpha-1-adrenergic receptor blockers, for examplefor use with darusentan, include carvedilol, doxazosin, labetalol,prazosin, terazosin and combinations thereof. It is noted that, ofthese, carvedilol and labetalol also function as beta-adrenergicreceptor blockers.

A suitable central alpha-2-adrenergic receptor agonist or othercentrally acting antihypertensive drug can illustratively be selectedfrom the following list:

clonidine

guanabenz

guanadrel

guanfacine

methyldopa

moxonidine

reserpine

A suitable aldosterone receptor antagonist can illustratively beselected from the following list:

canrenone

eplerenone

spironolactone

Still further classes of drugs that can be useful in combination oradjunctive therapy with darusentan or in a baseline antihypertensivetherapy include vasopeptidase inhibitors, NEP (neutral endopeptidase)inhibitors, prostanoids (particularly oral prostanoids), PDE5(phosphodiesterase type 5) inhibitors, nitrosylated compounds and oralnitrates.

Illustrative vasopeptidase inhibitors include:

fasidotril

omapatrilat

sampatrilat

Illustrative NEP inhibitors, some of which are also ACE inhibitors,include:

candoxatril

CGS 26582

MDL 100173

omapatrilat

phosphoramidon

sinorphan

thiorphan

Z13752A

Illustrative prostanoids include:

beraprost

cicaprost

epoprostenol

iloprost

PGE₁

PGI₂ (pro stacyclin)

NS-304

treprostinil

Illustrative PDE5 inhibitors include:

sildenafil

tadalafil

vardenafil

Other drugs that can be useful in combination or adjunctive therapy withdarusentan or in a baseline antihypertensive therapy can illustrativelybe selected from the following unclassified list:

ajmaline

alfuzosin

Alteon ALT 711

γ-aminobutyric acid

atrial natriuretic peptide

azelnidipine

bethanidine

bietaserpine

bosentan

budralazine

bufeniode

bunazosin

cadralazine

carmoxirole

CD 3400

chlorisondamine chloride

cicletanine

ciclosidomine

clevidipine

debrisoquin

denitronipradilol

desacetylalacepril

deserpidine

diazoxide

dihydralazine

endralazine

fenoldopam

flosequinan

guanethidine

guanidine, N-cyano-N′-4-pyridinyl-N″-(1,2,2-trimethylpropyl)-,monohydrate

guanoxabenz

guanoxan

hexamethonium

ketanserin

LBI 45

levcromakalim

lofexidine

magnesiocard

mebutamate

mecamylamine

normopresil

2-oxazolamine, N-(dicyclopropylmethyl)-4,5-dihydro-, (2E)-2-butenedioate

pargyline

pempidine

pentamethonium bromide

pentolinium tartrate

pheniprazine

phentolamine

pildralazine

pinacidil

piperoxan

protoveratrines

3,5-pyridinedicarboxylic acid,1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-, methyl1-(phenylmethyl)-3-pyrrolidinyl ester

raubasine

rescimetol

rescinnamine

rilmenidine

saralasin

sodium niroprusside

syrosingopine

Takeda TAK 536

TBC 3711

tetrahydrolipstatin

1,4-thiazepine-4(5H)-acetic acid,6-[[1-(ethoxycarbonyl)-3-phenylpropyl]amino]-tetrahydro-5-oxo-2-(2-thienyl)

tiamenidine

todralazine

tolonidine

trimethaphan camsylate

tyrosinase

urapidil

zofenopril

In one embodiment, the selective ET_(A) receptor antagonist, morespecifically darusentan, is administered concomitantly (e.g., incombination or adjunctive therapy) with one or more of

-   -   (a) a diuretic selected from the group consisting of        chlorothiazide, chlorthalidone, hydrochlorothiazide, indapamide,        metolazone, polythiazide, bumetanide, furosemide, torsemide and        combinations thereof;    -   (b) an ACE inhibitor selected from the group consisting of        benazepril, captopril, enalapril, fosinopril, lisinopril,        moexipril, perindopril, quinapril, ramipril, trandolapril and        combinations thereof, and/or an angiotensin II receptor blocker        selected from the group consisting of candesartan, eprosartan,        irbesartan, losartan, olmesartan, tasosartan, telmisartan,        valsartan and combinations thereof;    -   (c) a beta-adrenergic receptor blocker selected from the group        consisting of acebutolol, atenolol, betaxolol, bisoprolol,        carvedilol, labetalol, metoprolol, nadolol, penbutolol,        pindolol, propranolol, timolol and combinations thereof;    -   (d) a calcium channel blocker selected from the group consisting        of amlodipine, diltiazem, felodipine, isradipine, nicardipine,        nifedipine, nisoldipine, verapamil and combinations thereof;    -   (e) a direct vasodilator selected from the group consisting of        hydralazine, minoxidil and combinations thereof;    -   (f) an alpha-1-adrenergic receptor blocker selected from the        group consisting of carvedilol, doxazosin, labetalol, prazosin,        terazosin and combinations thereof;    -   (g) a central alpha-2-adrenergic receptor agonist or other        centrally acting drug selected from the group consisting of        clonidine, guanabenz, guanadrel, guanfacine, methyldopa,        moxonidine, reserpine and combinations thereof; and    -   (h) an aldosterone receptor antagonist selected from the group        consisting of canrenone, eplerenone, spironolactone and        combinations thereof.

More particularly, the selective ET_(A) receptor antagonist, morespecifically darusentan, can be administered in combination oradjunctive therapy with one or more of (a), (b), (c) and (d) above,optionally further with one or more of (e), (f), (g) and (h).

Still more particularly, the selective ET_(A) receptor antagonist, morespecifically darusentan, can be administered in combination oradjunctive therapy at least with (a) and any two of (b), (c) and (d).

As in the case of the selective ET_(A) receptor antagonist, the one ormore drugs constituting the baseline antihypertensive therapy andoptionally administered in combination with the selective ET_(A)receptor antagonist can be delivered by any suitable route ofadministration. Generally, such drugs are suitable for oraladministration, and many are suitable for once a day oraladministration. Thus in one embodiment at least one of the diuretic orantihypertensive drugs in the baseline therapy is orally administeredonce a day. In a particular embodiment, all drugs in the baselinetherapy are orally administered once a day. According to thisembodiment, it will generally be found convenient to administer alldrugs in the regimen, i.e., the selective ET_(A) receptor antagonist aswell as the baseline therapy drugs, orally once a day.

Fixed-dose combinations of two or more drugs can be achieved in manycases by coformulation of the drugs in a single dosage unit such as atablet or capsule. For example, coformulations of various drugs usefulin a baseline antihypertensive therapy as defined herein are available,including:

-   -   amiloride+hydrochlorothiazide;    -   amlodipine+benazepril;    -   atenolol+chlorthalidone;    -   benazepril+hydrochlorothiazide;    -   bisoprolol+hydrochlorothiazide;    -   candesartan+hydrochlorothiazide;    -   captopril+hydrochlorothiazide;    -   enalapril+felodipine;    -   enalapril+hydrochlorothiazide;    -   eprosartan+hydrochlorothiazide;    -   fosinopril+hydrochlorothiazide;    -   irbesartan+hydrochlorothiazide;    -   lisinopril+hydrochlorothiazide;    -   losartan+hydrochlorothiazide;    -   methyldopa+hydrochlorothiazide;    -   metoprolol+hydrochlorothiazide;    -   moexipril+hydrochlorothiazide;    -   nadolol+hydrochlorothiazide;    -   olmesartan+hydrochlorothiazide;    -   propranolol+hydrochlorothiazide;    -   quinapril+hydrochlorothiazide;    -   reserpine+chlorothiazide;    -   reserpine+chlorthalidone;    -   reserpine+hydrochlorothiazide;    -   spironolactone+hydrochlorothiazide;    -   telmisartan+hydrochlorothiazide;    -   timolol+hydrochlorothiazide;    -   trandolapril+verapamil;    -   triamterene+hydrochlorothiazide; and    -   valsartan+hydrochlorothiazide.

It will be understood that combination or adjunctive therapies asindicated above, while of particular benefit in patients havingresistant hypertension, are not limited to such patients. Hypertension(whether resistant or not) is a common feature of most complications ofdiabetes, including both diabetic nephropathy and metabolic syndrome,and it is contemplated that combination or adjunctive therapies of aselective ET_(A) receptor antagonist with one or more antihypertensivedrugs other than selective ET_(A) receptor antagonists can be useful inmany diabetic patients having these complications.

Another kind of combination or adjunctive therapy that can be usefulaccording to the present invention includes a selective ET_(A) receptorantagonist and one or more additional antidiabetic agents other thanselective ET_(A) receptor antagonists. Such additional antidiabeticagents can, for example, be selected from alpha-glucosidase inhibitors,biguanides, exendins, hormones and analogs thereof, meglitinides,sulfonylurea derivatives and thiazolidinediones.

A suitable antidiabetic can illustratively be selected from thefollowing list:

Biguanides

buformin

metformin

phenformin

Hormones and Analogs Thereof

amylin

insulin

insulin aspart

insulin detemir

insulin glargine

insulin glulisine

insulin lispro

liraglutide

pramlintide

Sulfonylurea Derivatives

acetohexamide

carbutamide

chlorpropamide

glibornuride

gliclazide

glimepiride

glipizide

gliquidone

glisoxepid

glyburide

glybuthiazole

glybuzole

glyhexamide

glymidine

tolazamide

tolbutamide

tolcyclamide

Thiazolidinediones

pioglitazone

rosiglitazone

troglitazone

Unclassified

acarbose

exenatide

miglitol

mitiglinide

muraglitazar

nateglinide

repaglinide

sitagliptin

tesaglitazar

vildagliptin

voglibose

Particularly suitable antidiabetics, for example for use withdarusentan, include acarbose, exenatide, glimepiride, insulins,metformin, nateglinide, pioglitazone, pramlintide, rosiglitazone andcombinations thereof.

For a subject with metabolic syndrome, a selective ET_(A) receptorantagonist can be administered in combination or adjunctive therapy withone or more additional agents selected from antidiabetics (for exampleas listed above), antihypertensives (for example as listed above),anti-obesity agents and antidyslipidemics.

Suitable anti-obesity agents include anorexics, CB1 receptor blockersand lipase inhibitors.

A suitable anti-obesity agent can illustratively be selected from thefollowing list:

aminorex

amphetamine

benzphetamine

chlorphentermine

clobenzorex

clortermine

cyclexedrine

dextroamphetamine

diethylpropion

N-ethylamphetamine

fenbutrazate

fenfluramine

fenproporex

levophacetoperane

mazindol

mefenorex

methamphetamine

norpseudoephedrine

orlistat

pentorex

phendimetrazine

phenmetrazine

phentermine

phenylpropanolamine

rimonabant

sibutramine

Particularly suitable anti-obesity agents, for example for use withdarusentan, include benzphetamine, methamphetamine, orlistat,phendimetrazine, phentermine, rimonabant, sibutramine and combinationsthereof.

Suitable antidyslipidemics include bile acid sequestrants, cholesterolabsorption inhibitors, fibrates, HMG CoA reductase inhibitors (statins),nicotinic acid derivatives, and thyroid hormones and analogs thereof.

A suitable antidyslipidemic can illustratively be selected from thefollowing list:

Bile Acid Sequestrants

cholestyramine resin

colesevelam

colestilan

colestipol

polidexide

Fibrates

bezafibrate

binifibrate

ciprofibrate

clinofibrate

clofibrate

clofibric acid

etofibrate

fenofibrate

gemfibrozil

pirifibrate

ronifibrate

simfibrate

theofibrate

HMG CoA Reductase Inhibitors

atorvastatin

cerivastatin

fluvastatin

lovastatin

pitavastatin

pravastatin

rosuvastatin

simvastatin

Nicotinic Acid Derivatives

acipimox

aluminum nicotinate

niacin (nicotinic acid)

niceritrol

oxiniacic acid

Thyroid Hormones and Analogs Thereof

dextrothyroxine

etiroxate

thyropropic acid

Unclassified

acifran

avasimibe

benfluorex

detaxtran

eicosapentaenoic acid

ezetimibe

meglutol

melinamide

omega-3 acid ethyl esters

γ-oryzanol

pantethine

pirozadil

policonasol

probucol

β-sitosterol

sultosilic acid

tiadenol

torcetrapib

xenbucin

Particularly suitable antidyslipidemics, for example for use withdarusentan, include atorvastatin, colesevelam, ezetimibe, fenofibrate,fluvastatin, lovastatin, niacin, rosuvastatin, simvastatin andcombinations thereof.

It is further contemplated that the selective ET_(A) receptor antagonistcan itself have useful antidyslipidemic activity, for example secondaryto its activity in enhancing glycemic control and/or insulinsensitivity.

When a selective ET_(A) receptor antagonist is used in adjunctivetherapy with one or more additional antidiabetics, antihypertensives,anti-obesity agents and/or antidyslipidemics, the selective ET_(A)receptor antagonist and at least one additional antidiabetic,antihypertensive, anti-obesity agent and/or antidyslipidemic can beadministered at different times or at about the same time (at exactlythe same time or directly one after the other in any order). Theselective ET_(A) receptor antagonist and the at least one antidiabetic,antihypertensive, anti-obesity agent and/or antidyslipidemic can beformulated in one dosage form as a fixed-dose combination foradministration at the same time, or in two or more separate dosage formsfor administration at the same or different times.

Separate dosage forms can optionally be co-packaged, for example in asingle container or in a plurality of containers within a single outerpackage, or co-presented in separate packaging (“common presentation”).As an example of co-packaging or common presentation, a kit iscontemplated comprising, in separate containers, darusentan and at leastone drug useful in combination or adjunctive therapy with darusentan,for example an antidiabetic, antihypertensive, anti-obesity agent orantidyslipidemic. In another example, darusentan and at least one druguseful in combination or adjunctive therapy with darusentan, for examplean antidiabetic, antihypertensive, anti-obesity agent orantidyslipidemic, are separately packaged and available for saleindependently of one another, but are co-marketed or co-promoted for useaccording to the invention. The separate dosage forms can also bepresented to a patient separately and independently, for use accordingto the invention.

A further embodiment of the present invention provides a method fortreating a complex of comorbidities in an elderly diabetic humansubject. This method comprises administering to the subject a selectiveET_(A) receptor antagonist in combination or adjunctive therapy with

-   -   (a) at least one additional agent that is (i) other than a        selective ET_(A) receptor antagonist and (ii) effective in        treatment of diabetes and/or at least one of said comorbidities        other than hypertension, and optionally    -   (b) at least one antihypertensive other than a selective ET_(A)        receptor antagonist.

An “elderly” subject is as defined hereinabove.

A “comorbidity” is a disease condition present in the subject inaddition to diabetes, that adds to the deleterious effects of thediabetes on the subject and/or affects the choice of therapy.Comorbidities can arise secondarily from the diabetes or from othercomorbidities, or may arise independently. Among comorbidities commonlyoccurring in an elderly diabetic patient population are, illustratively,insulin resistance, chronic kidney disease, hypertension, dyslipidemia,obesity, cardiac insufficiency and sleep apnea.

A “complex” of comorbidities is defined herein as the presence of atleast two such comorbidities, in addition to the underlying diabetes. Insome embodiments the subject presents with at least three, or even fouror more, such comorbidities. For example, in metabolic syndrome, asubject can exhibit diabetes with insulin resistance, hypertension,dyslipidemia and obesity.

“Treating” in the present context includes alleviating symptoms,enhancing glycemic control and/or insulin sensitivity, arresting,slowing, retarding or stabilizing progression of a condition or aphysiological or morphological marker thereof, and/or improving clinicaloutcome, for example as measured by quality of life, incidence orseverity of adverse cardiac events, time to end-stage renal disease orsurvival time.

The selective ET_(A) receptor antagonist can illustratively be selectedfrom those mentioned hereinabove. In one embodiment the selective ET_(A)receptor antagonist comprises darusentan, for example at dosage amountsand frequencies of administration, by routes of administration anddosage forms, and for duration of treatment as indicated hereinabove.

Several of the comorbidities mentioned above as occurring in elderlydiabetic patients have been individually reported in the literature(including literature cited herein) to be mediated by ET-1 and/or to betreatable with a selective ET_(A) receptor antagonist. However, it hasnot hitherto been proposed to simultaneously address a complex ofcomorbidities of diabetes in an elderly patient by treatment with aselective ET_(A) receptor antagonist such as darusentan; nor has it beenproposed to combine such treatment, in adjunctive or combinationtherapy, with one or more additional agents effective in treatment ofdiabetes and/or at least one of said comorbidities other thanhypertension. Given the spectrum of effects of selective ET_(A) receptorantagonists but without being bound by theory, it is believed that theselective ET_(A) receptor antagonist (e.g., darusentan) component ofsuch adjunctive or combination therapy can contribute in a substantialway to clinical improvement in each of a plurality of comorbidities, forexample supplementing the effect of, co-acting with, or permitting dosereduction (with potential benefits in reducing adverse side-effects) inat least one additional agent. However, even without such contributionby the selective ET_(A) receptor antagonist to more than onecomorbidity, the adjunctive or combination therapy of the presentembodiment brings great benefit to the elderly diabetic patient byenabling a complex of comorbidities, as seen for example in diabeticnephropathy or metabolic syndrome, to be simultaneously addressed.

The at least one additional agent that is (i) other than a selectiveET_(A) receptor antagonist and (ii) effective in treatment of diabetesand/or at least one comorbidity other than hypertension can comprise,for example, one or more antidiabetics, anti-obesity agents and/orantidyslipidemics, including any such agents listed hereinabove. Forexample, one or more agents selected from acarbose, exenatide,glimepiride, insulins, metformin, nateglinide, pioglitazone,pramlintide, rosiglitazone, benzphetamine, methamphetamine, orlistat,phendimetrazine, phentermine, rimonabant, sibutramine, atorvastatin,colesevelam, ezetimibe, fenofibrate, fluvastatin, lovastatin, niacin,rosuvastatin, simvastatin and combinations thereof can be administeredin adjunctive or combination therapy with a selective ET_(A) receptorantagonist, for example darusentan.

Antihypertensive(s) optionally additionally present in the adjunctive orcombination therapy can comprise, for example, agents of any classlisted hereinabove, including diuretics, ACE inhibitors, angiotensin IIreceptor blockers, beta-adrenergic receptor blockers, calcium channelblockers, direct vasodilators, alpha-1-adrenergic receptor blockers,central alpha-2-adrenergic receptor agonists and other centrally actingantihypertensive drugs, aldosterone receptor antagonists, vasopeptidaseinhibitors, NEP inhibitors, prostanoids, PDE5 inhibitors, nitrosylatedcompounds, oral nitrates and renin inhibitors, or combinations of agentsfrom one or more than one such class. For example, where the subject hasclinically diagnosed resistant hypertension as one of the comorbidities,the adjunctive or combination therapy can comprise administration of atleast one diuretic and at least two antihypertensives selected from atleast two of (a) ACE inhibitors and angiotensin II receptor blockers,(b) beta-adrenergic receptor blockers and (c) calcium channel blockers.

Illustratively antihypertensives for use in the method of the presentembodiment can be selected from chlorothiazide, chlorthalidone,hydrochlorothiazide, indapamide, metolazone, polythiazide, bumetanide,furosemide, torsemide, benazepril, captopril, enalapril, fosinopril,lisinopril, moexipril, perindopril, quinapril, ramipril, trandolapril,candesartan, eprosartan, irbesartan, losartan, olmesartan, tasosartan,telmisartan, valsartan, acebutolol, atenolol, betaxolol, bisoprolol,carvedilol, labetalol, metoprolol, nadolol, penbutolol, pindolol,propranolol, timolol, amlodipine, diltiazem, felodipine, isradipine,nicardipine, nifedipine, nisoldipine, verapamil and combinationsthereof.

A still further embodiment of the present invention provides atherapeutic combination comprising a selective ET_(A) receptorantagonist and at least one antidiabetic, anti-obesity orantidyslipidemic agent other than a selective ET_(A) receptorantagonist. Such a combination can have utility in a number ofsituations, not limited to methods described herein. However, acombination of this embodiment can be especially useful for treating acomplex of comorbidities in an elderly diabetic human subject asdescribed above.

The selective ET_(A) receptor antagonist and the at least oneantidiabetic, anti-obesity or antidyslipidemic agent can be present inthe combination in two or more separate dosage forms, permittingadministration at the same or different times. Such separate dosageforms can be formulated with one or more pharmaceutically acceptableexcipients for administration via the same or different routes. In aparticular embodiment all agents in the combination are formulated fororal administration; in an even more particular embodiment all agentsare formulated for once-daily oral administration and can beadministered at the same time each day. Where a treatment regimenincludes administration of a plurality of drugs, as in the presentinstance, there are great benefits in convenience and compliance instandardizing route, frequency and timing of administration in this way.

As indicated hereinabove, separate dosage forms can optionally beco-packaged, for example in a single container or in a plurality ofcontainers within a single outer package, or co-presented in separatepackaging, for example as a kit comprising, in separate containers, aselective ET_(A) receptor antagonist and at least one antidiabetic,anti-obesity or antidyslipidemic agent. The kit can optionally compriseseparate labeling information for each agent, or a single product labelhaving information on the therapeutic combination as a whole. In anotherexample, the selective ET_(A) receptor antagonist and the at least oneantidiabetic, anti-obesity or antidyslipidemic agent are separatelypackaged and available for sale independently of one another, but areco-marketed or co-promoted for use according to the invention. Theseparate dosage forms can also be presented to a patient separately andindependently, for use according to the invention.

In another embodiment, the selective ET_(A) receptor antagonist and theat least one antidiabetic, anti-obesity or antidyslipidemic agent arecoformulated with one or more pharmaceutically acceptable excipients ina single pharmaceutical composition as a fixed-dose combination.

A pharmaceutical composition comprising the selective ET_(A) receptorantagonist, the at least one antidiabetic, anti-obesity orantidyslipidemic agent, and one or more pharmaceutically acceptableexcipients is itself a still further embodiment of the presentinvention.

In a therapeutic combination or pharmaceutical composition of theinvention, the selective ET_(A) receptor antagonist can illustrativelybe selected from those mentioned hereinabove. In one embodiment theselective ET_(A) receptor antagonist comprises darusentan, for examplein a dosage amount as set forth hereinabove.

The combination or composition can comprise at least one antidiabetic,for example selected from alpha-glucosidase inhibitors, biguanides,exendins, hormones and analogs thereof, meglitinides, sulfonylureaderivatives and thiazolidinediones. In a particular embodiment theselective ET_(A) receptor antagonist comprises darusentan and the atleast one antidiabetic is selected from acarbose, exenatide,glimepiride, insulins, metformin, nateglinide, pioglitazone,pramlintide, rosiglitazone and combinations thereof.

The combination or composition can comprise at least one anti-obesityagent, for example selected from anorexics, CB1 receptor blockers andlipase inhibitors. In a particular embodiment the selective ET_(A)receptor antagonist comprises darusentan and the at least oneanti-obesity agent is selected from benzphetamine, methamphetamine,orlistat, phendimetrazine, phentermine, rimonabant, sibutramine andcombinations thereof.

The combination or composition can comprise at least oneantidyslipidemic, for example selected from bile acid sequestrants,cholesterol absorption inhibitors, fibrates, HMG CoA reductaseinhibitors, nicotinic acid derivatives, and thyroid hormones and analogsthereof. In a particular embodiment the selective ET_(A) receptorantagonist comprises darusentan and the at least one antidyslipidemic isselected from atorvastatin, colesevelam, ezetimibe, fenofibrate,fluvastatin, lovastatin, niacin, rosuvastatin, simvastatin andcombinations thereof. For example, the combination or composition cancomprise a selective ET_(A) receptor antagonist such as darusentan, acholesterol absorption inhibitor such as ezetimibe, and an HMG CoAreductase inhibitor (statin) such as atorvastatin, fluvastatin,lovastatin, rosuvastatin or simvastatin.

The combination or composition comprises, in one embodiment, a selectiveET_(A) receptor antagonist, at least one antidiabetic and at least oneanti-obesity agent.

The combination or composition comprises, in another embodiment, aselective ET_(A) receptor antagonist, at least one antidiabetic and atleast one antidyslipidemic.

The combination or composition comprises, in yet another embodiment, aselective ET_(A) receptor antagonist, at least one anti-obesity agentand at least one antidyslipidemic.

The combination or composition comprises, in a still further embodiment,a selective ET_(A) receptor antagonist, at least one antidiabetic, atleast one anti-obesity agent and at least one antidyslipidemic.

According to any embodiment mentioned above, the combination orcomposition optionally further comprises at least one antihypertensive.The at least one antihypertensive can comprise, for example, one or moreagents of any class listed hereinabove or a combination of agents frommore than one such class.

An illustrative combination or composition of the invention comprises:

-   -   (a) darusentan as a selective ET_(A) receptor antagonist;    -   (b) one or more of:        -   (i) at least one antidiabetic selected from acarbose,            exenatide, glimepiride, insulins, metformin, nateglinide,            pioglitazone, pramlintide, rosiglitazone and combinations            thereof;        -   (ii) at least one anti-obesity agent selected from            benzphetamine, methamphetamine, orlistat, phendimetrazine,            phentermine, rimonabant, sibutramine and combinations            thereof; and/or        -   (iii) at least one antidyslipidemic selected from            atorvastatin, colesevelam, ezetimibe, fenofibrate,            fluvastatin, lovastatin, niacin, rosuvastatin, simvastatin            and combinations thereof; and    -   (c) at least one antihypertensive selected from chlorothiazide,        chlorthalidone, hydrochlorothiazide, indapamide, metolazone,        polythiazide, bumetanide, furosemide, torsemide, benazepril,        captopril, enalapril, fosinopril, lisinopril, moexipril,        perindopril, quinapril, ramipril, trandolapril, candesartan,        eprosartan, irbesartan, losartan, olmesartan, tasosartan,        telmisartan, valsartan, acebutolol, atenolol, betaxolol,        bisoprolol, carvedilol, labetalol, metoprolol, nadolol,        penbutolol, pindolol, propranolol, timolol, amlodipine,        diltiazem, felodipine, isradipine, nicardipine, nifedipine,        nisoldipine, verapamil and combinations thereof.

EXAMPLES

The following examples are merely illustrative, and do not limit thisdisclosure in any way. Reference is made in the examples to statisticalanalysis. Such reference is made in the interest of full disclosure anddoes not constitute admission that statistical significance is aprerequisite for patentability of any claim herein.

Example 1

Summary

The following is a protocol for a Phase II double blindplacebo-controlled randomized study to investigate the safety and thehemodynamic effects of three-week oral applications of different dosesof darusentan on top of standard medication in congestive heart failurepatients.

Objectives

The primary objective of this study was to assess the tolerability andsafety profile of three-week oral applications of different doses ofdarusentan on top of established standard medicinal treatments inpatients suffering from advanced chronic congestive heart failure NYHAfunctional class III. Secondary objective was the assessment of theshort-term effects of three-week darusentan treatment on hemodynamicparameters by means of SWAN GANZ floating catheterization andthermodilution.

Methodology

This was a multinational, multicenter, prospective, double-blind,placebo-controlled, randomized clinical trial.

After a pre-investigational eligibility check, double-blind treatmentwas started for a period of three weeks, followed by a one-weekdouble-blind follow-up period.

The safety and hemodynamic effects of three different dosages ofdarusentan were consecutively studied, starting with darusentan at 30 mgor placebo. The following dosages of 300 mg followed by 100 mg wereselected after reviewing the results and experiences gleaned fromevaluating the safety data of at least 15 patients who completed theentire three-week dosing period.

Patients

To obtain at least 24 evaluable patients per treatment group, it wasexpected to include a total number of 120 patients into the study.Actually, 157 patients were randomized (30 mg: n=36; 100 mg: n=39; 300mg: n=49; placebo: n=33).

Male and female patients were selected based on the following profile:aged 18 years or older who needed to undergo SWAN GANZ floatingcatheterization for diagnostic reasons with clinical CHF signs of atleast three months duration and for any underlying cause (except forprimary organic valvular heart disease), present or recent history ofNYHA functional class III and presenting with a left ventricularejection fraction less than or equal to 35%, assessed by means ofechocardiography or isotope ventriculography within seven days prior toinvestigational procedures. Continuation in the study was only possible,if at baseline pulmonary capillary wedge pressure (PCWP) was more thanequal to 12 mmHg and cardiac index (CI) was less than or equal to 2.6l/min/m².

Test Product, Dose, Mode of Administration and Duration of Treatment

Tablets containing 30 mg, 100 mg or 300 mg darusentan.

Medication was taken orally once a day for 3 weeks

Reference Therapy, Dose, Mode of Administration and Duration ofTreatment

Matching placebo tablets taken orally once a day for 3 weeks.

Criteria for Evaluation:

1. Efficacy

Changes in hemodynamic parameters from baseline focusing on CI(l/min/m²), and PCWP (mmHg). Furthermore, pharmacokinetic parameters andneurohormone plasma levels were assessed.

2. Safety

Adverse events, changes in systemic blood pressure, heart rate, ECG andclinical laboratory parameters were measured.

Statistical Methods

All treated patients were included in the safety analysis. Efficacy wasanalyzed primarily according to the intention-to-treat principle.Continuous data were described by statistical characteristics (m, mean,standard deviation, minimum, 1^(st) quartile, median, 3^(rd) quartile,maximum, number of missing values) for each time point, as well as forchanges from baseline. For categorical data and adverse events,frequency and percentage were given. For the two primary efficacyparameters CI (l/min/m²) and PCWP (mmHg), analysis-of-covariance modelswere calculated.

Results:

During the course of this study blood glucose levels (mg/dl or mmol/l)were determined at patient visits. Only fasting blood glucose levels areshown below. “N/A” is used where either blood glucose level was notexamined or the laboratory test was performed when the patient was notfasting.

Glucose Levels Visit Visit Visit Visit Visit Visit 1 2 3 4 5 6 SideEffects Patient N/A 156 N/A N/A N/A 139 Increasing of 185 Dyspnoea (3)Increase of Body Weight(3) Decrease of Diuresis (3) Patient N/A 11.4 8.66.4 11.1 5.2 Hyperglycemia (5) 152 Face Rush (3) Pneumonia (3) Headacheeach day (2) Better Exercise Tolerance (2) Patient N/A 116 121 N/A N/AN/A Sensations of Heat 190 (3) Head cephalgia (3) Angina Pectoris (3)Subjective feelings of unrest (3) Sleep Disturbance (3) Angina Pectoriswith Dyspnoea(3) Sensations of Abdominal Pressure (3) Patient N/A 97 92N/A N/A N/A CHF worsening (4) 215 Patient N/A 127 141 N/A N/A 104 AcuteLeft Heart 216 Failure (5) Decreasing Plueral Effusion Patient 80 65 N/AN/A N/A 88 None Listed 288 Side Effect Relationship to Drug 1-Definite2-Probable 3-Possible 4-Unlikely 5-Unrelated **Non-survivor patientswhose mordity was deemed to have an unlikely or unrelated relationshipto darusentan by the clinician have been omitted.

Example 2

Summary

Hypertension clinical studies conducted over the last decade haveindicated that proper control of systolic blood pressure (SBP) isequally as important as diastolic blood pressure (DBP) control inrelationship to cardiovascular and renal outcomes; and SBP is moredifficult to control than DBP, especially in patients over 50 years ofage. Despite treatment with multiple antihypertensive drugs attherapeutic doses, a substantial number of patients, especially thosewith diabetes mellitus (diabetes) and/or chronic kidney disease (CKD),do not reach guideline-recommended blood pressure goals.

The following is the protocol for a phase 3 randomized, double-blind,placebo-controlled, multi-center, parallel group study to evaluate theefficacy and safety of fixed doses of darusentan in subjects withresistant systolic hypertension receiving combination therapy with threeor more antihypertensive drugs, including a diuretic.

LIST OF ABBREVIATIONS ABPM Ambulatory blood pressure monitoring IECIndependent ethics committee ACEI Angiotensin converting enzymeinhibitor IgM Immunoglobulin M AE Adverse event IND Investigational NewDrug application ALT Alanine aminotransferase INR Internationalnormalized ratio ANCOVA Analysis of covariance IRB Institutional reviewboard ARB Angiotensin receptor blocker kg Kilograms AST Aspartateaminotransferase K_(i) Inhibition constant AUC Area-under-the-curve LDHLactate dehydrogenase AV Atrioventricular LDL Low density lipoproteinBMI Body mass index LFT Liver function test BNP b-type natriureticpeptide LH Luteinizing hormone BP Blood pressure LLN Lower limit ofnormal BUN Blood urea nitrogen LV Left ventricular CCB Calcium channelblocker LVIDD Left ventricular internal diastolic diameter C_(max) Peakplasma concentration m² Meters squared CHF Chronic heart failure MDRDModification of Diet in Renal Disease CFR Code of Federal Regulations mgMilligrams CKD Chronic kidney disease min Minutes CNO Certificate ofnon-objection mL Milliliters CRF Case report form mmHg Millimeters ofmercury DBP Diastolic blood pressure μg Micrograms dL Deciliters nMNanomolar DMC Data Monitoring Committee NOTEL No toxic effect level ECGElectrocardiogram NOAEL No observed adverse effect level eGFR Estimatedglomerular filtration rate NSAIDs Non-steroidal anti-inflammatory drugsET-1 Endothelin-1 P450 Cytochrome P450 ET_(A) Endothelin A receptor PDCPremature Discontinuation ET_(B) Endothelin B receptor PDEPhosphodiesterase ERA Endothelin receptor antagonist PHI Protectedhealth information FDA Food and Drug Administration PK PharmacokineticFSH Follicle stimulating hormone po Per os (orally) g Grams PTProthrombin time GCP Good clinical practice PTT Partial thromboplastintime GDC Global Data Collection qd Once daily GGT Gamma glutamylaminotransferase RBC Red blood cell HbA_(1c) Glycosylated hemoglobinRHTN Resistant hypertension HDL High density lipoprotein SADR Seriousadverse drug reaction HIPAA Health Insurance Portability andAccountability Act SAE Serious adverse event HR Heart rate SaO₂ Arterialoxygen saturation IB Investigator's Brochure SBP Systolic blood pressureICD Implantable cardioverter defibrillator SCr Serum creatinine ICFInformed Consent Form SSRIs Selective serotonin reuptake inhibitors ICHInternational Conference on Harmonization SUSAR Suspected unexpectedserious adverse event t_(1/2) Elimination half-life TCA Tricyclicantidepressants TG Triglycerides TSH Thyroid stimulating hormone UACRUrinary albumin-to-creatinine ratio ULN Upper limit of normal WBC Whiteblood cell

METHODS

Patients

Approximately 352 subjects will be randomized to one of three doses ofdarusentan (50, 100 or 300 mg po qd) or placebo in a ratio of 7:7:7:11at approximately 160 investigative sites in North and South America,Europe, New Zealand, and Australia.

Eligible subjects will include men and women, 35-80 years old, treatedwith full doses of three or more antihypertensive drugs, including adiuretic, with RHTN as defined by contemporary clinical guidelines forthe treatment of hypertension [1,2]. Subjects with diabetes and/or CKDmust have a SBP ≧130 mmHg at Screening to be eligible for study entry.All other subjects must have a SBP of ≧140 mmHg. SBP at Screening mustbe <180 mmHg for all subjects. A BMI of 20 to 43 kg/m² or an upper armcircumference <42 cm, and an eGFR ≧30 mL/min/1.73 m² at Screening arealso required. Subject eligibility will be reassessed at the initiationof the 2-week single-blind Placebo Run-in Period and at theRandomization Visit (see Study Schematic), and only those subjects whocontinue to meet inclusion/exclusion criteria at these visits will berandomized into the study. In particular, SBP and DBP measured at thePlacebo Run-in Visit and at the Randomization Visit must be within 20mmHg and 10 mmHg, respectively, of the values recorded at Screening inorder for the subject to be eligible for randomization. NOTE: Subjectswho fail to meet entry criteria after entering the placebo run-in phasewill not be allowed to re-screen for the study.

All potential subjects will be classified with regard to diabetes andCKD as part of the Screening process. Diabetic subjects must have adocumented diagnosis of Type 2 diabetes prior to Screening, and allscreened subjects will be evaluated for the presence of CKD according tothe following definition: (i) reduced excretory function with an eGFR<60 mL/min/1.73 m² and/or (ii) the presence of albuminuria in a spoturine sample (>200 mg/g [22.60 mg/mmol] creatinine).

Antihypertensive therapy must include:

-   -   A diuretic, preferably a thiazide; and    -   Two or more drugs from at least two of the following classes of        antihypertensive agents:        -   ACEIs, ARBs, and/or renin inhibitors        -   CCBs        -   beta-blockers        -   central alpha-2 agonists        -   peripheral alpha-1 antagonists        -   direct vasodilators        -   other centrally-acting drugs

The combination of antihypertensive drugs administered should beconsistent with current treatment guidelines [1,2]. For example, commonmulti-drug therapy may include a thiazide diuretic, an ACEI or an ARB,and a CCB. Subjects may be on more than 3 antihypertensive drugs atentry as long as the minimum requirements described above are met.

The minimum allowable dose of hydrochlorothiazide (HCTZ) will be 25 mg.For other thiazide-type diuretics, an equivalent dose to 25 mg HCTZ isalso required. A loop diuretic may be substituted for the thiazidediuretic in subjects with CKD or a documentedcontraindication/intolerance to treatment with a thiazide. Apotassium-sparing diuretic alone (e.g., triamterene) will not qualify asadequate diuretic therapy. The dose of each antihypertensive medicationthat the subject is receiving will be documented at Screening, andmonitored throughout study participation. In addition, the dose of eachconcomitant antihypertensive medication will be classified at studyentry according to the following criteria:

-   -   Highest labeled dose according to the product's package        insert/labeling information for the applicable country/region    -   Highest usual dose per clinical guidelines [1]    -   Highest tolerated dose    -   Highest appropriate dose for the subject per the Principal        Investigator's best clinical judgment

It is expected that subjects will have been optimized on theirantihypertensive drug regimens well in advance of Screening for thisstudy. Subjects must be stable on all antihypertensive drugs for atleast 4 weeks prior to the Screening Visit. Adjustments to the number ordosage of concomitant antihypertensive medications will not be permittedat any time during the study, with the exception of protocol-allowedchanges to diuretic therapy implemented to specifically address fluidretention-related events (see Study Design section below).

Subjects who meet any one of the following criteria will be deemedineligible for participation in the study:

-   1. Subjects with an average sitting SBP of ≧180 mmHg or DBP of ≧110    mmHg.-   2. Subjects with left ventricular (LV) systolic dysfunction as    evidenced by a LV ejection fraction <40% and/or a LV internal    diastolic diameter (LVIDD)>3.2 cm/m² or >6.0 cm, measured by    echocardiogram at Screening (Visit 1) or within 3 months prior to    Screening.-   3. Subjects with a HbA_(1c)>10% at Screening (Visit 1).-   4. Subjects who have:    -   a. A hemoglobin concentration <11.5 g/dL at Screening (Visit 1)        or    -   b. A hematocrit <34% at Screening (Visit 1)-   5. Subjects with hypo- or hyperthyroidism, as evidenced by a serum    thyroid stimulating hormone (TSH) concentration >1.5×ULN or <1.5×    the lower limit of normal (LLN) at Screening (Visit 1).-   6. Subjects with a serum ALT or AST >2×ULN at Screening (Visit 1).-   7. Subjects with other identifiable secondary causes of resistant    hypertension (e.g., parathyroid disease, pheochromocytoma, aortic    coarctation, Cushing's disease, hyperaldosteronism).-   8. Subjects who have experienced a myocardial infarction, unstable    angina pectoris, or a cerebrovascular accident within 6 months of    the Screening Visit (Visit 1).-   9. Subjects with sick sinus syndrome or second or third degree    atrioventricular (AV) block, chronic atrial fibrillation or    recurrent atrial tachyarrhythmia (including paroxysmal atrial    tachycardia), a history of recurrent ventricular tachycardia, or    symptomatic bradycardia.-   10. Subjects with implanted pacemakers or an implanted cardioverter    defibrillator (ICD).-   11. Subjects with a historical or current diagnosis of symptomatic    or asymptomatic CHF, treated or untreated.-   12. Subjects with hemodynamically significant valvular heart    disease.-   13. Subjects with Type 1 diabetes mellitus.-   14. Subjects on hemodialysis or peritoneal dialysis at the time of    Screening (Visit 1) and subjects with a history of solid organ    transplant (e.g., kidney, heart).-   15. Subjects who have had a diagnosis or recurrence of malignancy    within the past 3 years, with the exception of basal cell carcinoma    of the skin or in situ carcinoma of the cervix.-   16. Subjects with sleep apnea are excluded, unless a post-treatment    sleep study has confirmed treatment efficacy and there are no    recordings of blood oxygen saturation (SpO₂)<90% at any time during    the testing period.-   17. Subjects who perform alternating shift or night work.-   18. Women of childbearing potential or women who are pregnant or    nursing.-   19. Subjects not on stable doses of all concomitant medications for    a minimum of 4 weeks prior to Screening, and subjects treated with    any of the following prohibited medications:    -   a. Oral or injected corticosteroids within 3 months of Screening        (Visit 1). Systemic treatment with oral or injected steroids is        also prohibited during study participation. NOTE: The use of        inhaled steroids is allowed.    -   b. Aspirin in excess of 325 mg per day.    -   c. Chronic stable or unstable use of non-steroidal        anti-inflammatory drugs (NSAIDS) other than aspirin is        prohibited. Chronic use is defined as consecutive or        nonconsecutive days of treatment per week. (Note: In addition,        the intermittent use of NSAIDs is strongly discouraged        throughout the duration of this study. If intermittent treatment        is required, NSAIDs must not be used for more than a total of 2        days during the single-blind Placebo Run-in Period or the 2        weeks prior to the Week 14 Visit [Visit 10a]. For all subjects        requiring analgesic or anti-pyretic agents, the use of        acetaminophen is recommended during study participation).    -   d. Selective serotonin reuptake inhibitors (SSRIs), if a subject        is not compliant with the medication and/or has not been        receiving a stable dose for at least 3 months prior to        Screening. In addition, SSRIs or similar drugs for the treatment        of anxiety or depression may not be initiated at any time during        the study.    -   e. Tricyclic antidepressants (TCAs), if a subject is not        compliant with the medication and/or has not been receiving a        stable dose for at least 3 months prior to Screening. In        addition, TCAs or similar drugs for the treatment of anxiety or        depression may not be initiated at any time during the study.    -   f. Another ERA (e.g., bosentan, sitaxsentan, atrasentan,        TBC3711) at or within 6 months of Screening (Visit 1). In        addition, subjects may not initiate treatment with another ERA        at any time during the study.    -   g. The use of short-acting oral nitrates (e.g., sublingual        nitroglycerin) is permitted; however, subjects should not take        short-acting oral nitrates within 4 hours of Screening (Visit 1)        or any subsequent study visit.    -   h. The use of long-acting oral nitrates (e.g., Isordil) is        permitted; however, the dose must be stable for at least 2 weeks        prior to Screening (Visit 1) and Randomization (Visit 3a).    -   i. The use of oral sympathomimetic decongestants or β-agonists        is permitted; however, not within 1 week prior to Screening        (Visit 1), initiation of the Placebo Run-in (Visit 2), or        Randomization (Visit 3a). In addition, use of these medications        will be prohibited within 1 day prior to any clinic visit during        study participation. NOTE: The stable chronic use of inhaled        β-agonists is permitted. These drugs may be used as prescribed        without the visit-based prohibitions described above.    -   j. The use of theophylline is permitted; however, the dose must        be stable for at least 4 weeks prior to Screening (Visit 1) and        throughout study participation.    -   k. The use of phosphodiesterase (PDE) type V inhibitors is        permitted; however, subjects must refrain from taking these        medications within three (3) days of Screening (Visit 1) or any        subsequent study visit.    -   l. Use of thiazolidinedione (i.e., glitazone) class of        anti-diabetic medications (e.g., rosiglitazone, pioglitazone;        alone or in combination pills) at or within 4 weeks of Screening        (Visit 1). Subjects should not initiate treatment with        thiazolidinediones at any time during the study.    -   Note: Subjects receiving exclusionary medications at Screening        must undergo a minimum of 4 weeks washout prior to being        re-screened for the study.-   20. Subjects who have demonstrated non-compliance with previous    medical regimens.-   21. Subjects with a contraindication to treatment with an ERA.    Contraindications include, but are not limited to, a history of    elevated liver function tests (e.g., aminotransferases >2×ULN) or an    event defined as a serious adverse event (SAE) attributed to    previous treatment with an ERA.-   22. Subjects who participated in a prior clinical study of    darusentan and were randomized to, and received, active treatment.-   23. Subjects who have participated in a clinical study involving    another investigational drug or device within 4 weeks of the    Screening Visit (Visit 1).-   24. Subjects who have failed screening for this study two times.-   25. Subjects who have any concomitant condition that, in the opinion    of the investigator, may adversely affect the safety and/or efficacy    of the study drug or severely limit the subject's lifespan or    ability to complete the study (e.g., alcohol or drug abuse,    disabling or terminal illness, mental disorders,    institutionalization by judicial order).    Objectives

The primary objective of this study is to determine if darusentan iseffective in reducing SBP and DBP in subjects with RHTN, despitetreatment with full doses of three or more antihypertensive drugs,including a diuretic.

Secondary objectives of this study are to examine the effect ofdarusentan on mean 24-hour ambulatory blood pressure, percent ofsubjects meeting SBP goal, and eGFR. The safety and tolerability ofdarusentan in the subject population will also be evaluated.

Several measures of interest will also be examined.

Study Design

This is a Phase 3 randomized, double-blind, placebo-controlled,multi-center, parallel group study in subjects with RHTN, despitetreatment with full doses of three or more antihypertensive drugs,including a diuretic.

The study will consist of three periods: Screening, Placebo Run-in, andTreatment. Screening assessments and evaluations may be conducted over aperiod of not more than 2 weeks. Following Screening, all eligiblesubjects will undergo a single-blind, Placebo Run-in of 2 weeks durationto ensure that blood pressure remains stable and continues to meeteligibility criteria for randomization. Subjects who continue to meeteligibility criteria following the 2-week Placebo Run-in Period will berandomized to one of four treatment groups (50, 100, or 300 mg ofdarusentan or placebo po qd), stratified by co-morbidity status (i.e.,presence of diabetes and/or CKD versus the absence of these conditions)and race (i.e., Black versus non-Black). Subjects randomized to placeboor 50 mg darusentan will receive their randomized dose of study drugthroughout the 14-week Treatment Period. Subjects randomized to 100 or300 mg darusentan will be initiated on 50 mg for 2 weeks and willsubsequently undergo up-titration to the next higher dose of darusentanevery 2 weeks until the randomized dose is achieved (see FIG. 1). If asubject experiences a severe study drug-related AE during the TreatmentPeriod, the subject's study drug dose may be reduced, according toinvestigator discretion. The choice to reduce the study drug dose may bemade only once during the Treatment Period, and the change must beimplemented prior to or at the Week 6 Visit. Once a subject's study drugdose has been down-titrated, it may not be subsequently increased.Subjects requiring down-titration of study drug after study Week 6 mustdiscontinue treatment with study drug; however, the subject shouldremain in study through the end of the Treatment Period (i.e., Week 14).Study drug assignments will remain double-blinded throughout theTreatment Period. All subjects will receive a fixed dose of study drugfor a minimum of 8 weeks prior to the evaluation of study endpoints atthe Week 14 Visit.

Adjustments to the number or dosage of concomitant antihypertensivemedications will not be permitted at any time during the study. However,if a subject develops signs or symptoms of fluid retention during theTreatment Period, manifested as peripheral edema and/or clinicallysignificant weight gain between study visits that in the opinion of theinvestigator requires immediate treatment, the subject's diuretictherapy may be adjusted according to investigator discretion. It isrecommended that a loop diuretic be added, or the dose increased forsubjects already receiving a loop diuretic, prior to adjustingconcomitant thiazide diuretic therapy. All changes will be documented indetail in the subject's case report form (CRF). Adjustment of diureticswill not be allowed within 2 weeks of the primary endpoint assessment.

Subjects who complete participation in the study through the Week 14Visit, on or off study drug, will have the option to participate in along-term safety extension study, with the exception of subjects whodiscontinue study drug due to a study drug-related AE. Subjects whodiscontinue treatment with study drug prior to the end of the TreatmentPeriod due to a study drug-related AE will not be eligible toparticipate in the long-term safety extension study. Subjects who do notparticipate in the long-term safety extension study will discontinuetreatment with study drug at the Week 14 Visit, and will return to theclinic for two safety visits prior to concluding study participation.Maximum placebo exposure in this study will be 16 weeks.

Women of childbearing potential will be excluded from studyparticipation. Women who are surgically sterile or documentedpost-menopausal for at least 2 years are not considered to be ofchildbearing potential. Post-menopausal female subjects who are notsurgically sterile will be required to use a double-barrier method ofbirth control throughout study participation. Serum alanineaminotransferase (ALT), aspartate aminotransferase (AST), alkalinephosphatase, gamma glutamyl aminotransferase (GGT), and total bilirubinwill be monitored in all subjects throughout the study. All men will berequired to provide blood samples for hormone analyses, and men who areable will be required to undergo semen analyses prior to and duringtreatment with study drug in order to evaluate the potential effects ofdarusentan on male fertility. Men who have had a vasectomy or who areunable to provide semen samples will be excused from this requirement.

Schedule of Assessments

STUDY VISIT 1 2 3a 3b 4 5 6 7 8 9 10a 10b Study Week Random. 14 −4 −2 D1 D 2 2 4 6 8 10 12 D 1 D 2 UV¹ Assessments Informed consent X ReviewInclusion/ X X X Exclusion criteria Medical X history/demographicsConMed assessment X X X X X X X X X X X Adverse Event X X X X X X X X XX assessment Trough sitting/standing X X X X X X X X X X X BP and HRFull physical examination X X Abbreviated physical X X X X X X X X Xexamination Measure body weight X X X X X X X X X X X 12-lead ECG X X XX X Echocardiogram X² Ambulatory BP on off on off Monitoring HealthEconomics X³ Questionnaire Obtain vital status⁴ Laboratory TestsChemistry X⁵ X X X X X X Hematology X X X X X X X Urine sample X X X⁶ X⁶X⁶ X⁶ X X⁶ Fasting Blood Collection⁷ X, S S Trough PK plasma X X X Xsample Biomarker plasma X X sample Women only: Serum pregnancy test X Xβ-hCG urine pregnancy X test Men only: Hormone analysis X X Semenanalysis S^(8▪▪) ^(▪)(X) S^(8▪▪) ^(▪)(X) Warfarin-treated subjects only:Coagulation⁹ X X Study Drug Collect study drug/assess X X X X X X X XX¹⁰ compliance Register study visit X X X X X X X X X X X X XRandomization X Dispense study drug X¹¹ X¹² X X X X X X X¹⁰ ¹UV =Unscheduled Visit; ²An echocardiogram must be obtained during theScreening Period or within 3 months prior to Screening. However,historical echocardiograms must report predefined criteria necessary todetermine subject eligibility. ³A Health Economics Questionnaire will becompleted by all screened subjects. ⁴All randomized subjects must becontacted to assess vital status at or shortly after the time of thelast subject visit and again for the purposes of Global Data Collection.⁵The chemistry panel performed at Screening will include measurements ofimmunoglobulin M (IgM), thyroid stimulation hormone (TSH) andglycosylated hemoglobin (HbA_(1c)). TSH and HbA_(1c) will be used toevaluate subject eligibility. ⁶Spot urine collection for subjects withclinically significant albuminuria (≧30 mg/g [3.39 mg/mmol] creatinine)at Screening. ⁷S = Schedule. Fasting Blood Collection-Baseline may occurat Placebo Run-in (Visit 2) or within 2 weeks after that visit, but notat the Randomization Visit (Visit 3). Fasting Blood Collection-End ofStudy (EOS) for all subjects may occur within the 2 weeks following theWeek 12 Visit (Visit 9), but not during the Week 14 Visit (Visit 10).⁸Schedule a semen sample collection within 2 weeks prior to or duringRandomization (Visit 3a). Schedule a post-baseline semen samplecollection within 2 weeks prior to or during the Week 14 Visit (Visit10a). ⁹Coagulation labs will be completed for subjects receivingwarfarin (Coumadin) or warfarin-like anticoagulants. ¹⁰If down-titrationof study drug occurs at an unscheduled visit, collection of study drug,assessment of compliance, and dispensation of study drug should also beperformed. ¹¹Single-blind, placebo study drug will be dispensed to alleligible subjects at the visit. ¹²Randomized, double-blind treatmentbegins for all eligible subjects.Primary Efficacy Endpoints

The co-primary endpoints are the change from baseline to Week 14 intrough sitting SBP and trough sitting DBP, as measured bysphygmomanometry.

Secondary Endpoints

The following secondary endpoints will be assessed during this study:

-   -   Change from baseline to final measurement in mean 24-hour SBP        measured by ABPM    -   Change from baseline to final measurement in mean 24-hour DBP        measured by ABPM    -   The percent of subjects who reach SBP goal after 14 weeks of        treatment, defined as follows:        -   Subjects with diabetes and/or CKD must reach a SBP goal of            <130 mmHg        -   All other subjects must reach a SBP goal of <140 mmHg    -   Change from baseline to Week 14 in eGFR        Measures of Interest

The following measures of interest will be examined:

-   -   Change from baseline in the following ABPM measures:        -   Mean hourly ambulatory SBP and DBP over a 24-hour monitoring            period        -   Mean trough ambulatory SBP and DBP        -   Mean peak ambulatory SBP and DBP        -   Mean trough/peak ratio        -   Mean daytime ambulatory SBP and DBP        -   Mean nighttime ambulatory SBP and DBP        -   Mean daytime/nighttime ratio;    -   Change from baseline to Week 14 in pulse pressure measured by        sphygmomanometry;    -   Change from baseline in selected biomarker concentrations;    -   Change from baseline in glycosylated hemoglobin (HbA_(1c)),        fasting plasma glucose, serum lipid profile, serum insulin,        serum C-peptide, serum C-reactive protein, and waist        circumference;    -   Change from baseline in urinary albumin-to-creatinine ratio        (UACR) in a subset of subjects with clinically significant        albuminuria (i.e., ≧30 mg/g [3.39 mg/mmol] creatinine in a spot        urine sample);    -   Trough plasma concentrations of darusentan and its metabolites;        Statistical Methods

Change in SBP and DBP from baseline to Week 14 will be tested withanalysis of covariance, using stratification factors (comorbidity statusand race) and baseline blood pressure as covariates. Proportion at goalblood pressure will be tested with logistic regression, using the samecovariates. The last observation during double blind therapy will beused for analyses.

Adjustment for multiple comparisons for the primary endpoints of changein SBP and DBP will use the fallback method. First, SBP will be comparedbetween the 300 mg dose and placebo at the nominal alpha level of 0.04.If this is significant, DBP will be compared between the 300 mg dose andplacebo at the nominal level of 0.04. From this point, the fixedsequence procedure will be followed, requiring significance on anendpoint before allowing testing of the following endpoint, and testingall comparisons at the same level. If the 300 mg dose was significant atthe 0.04 level for both SBP and DBP, the tests will be reported at the0.05 level; otherwise the tests will be reported at the 0.01 level.These comparisons will be, in order, the 100 mg dose compared to placebofor SBP then DBP, and the 50 mg dose compared to placebo for SBP andthen DBP. Continuous secondary endpoints will be handled analogously.

Assuming a difference in placebo-adjusted change from baseline of 8 mmHgin trough SBP for each darusentan dose and a standard deviation of 15mmHg, 121 subjects randomized to placebo and 77 subjects randomized toeach dose of darusentan will provide 85-90% power to detect a differencebetween placebo and any individual darusentan dose, and 95% power tofind at least one dose of darusentan that is different from placebo.

It is expected that treatment of a human subject, for example a humansubject having diabetic nephropathy and/or metabolic syndrome, inaccordance with the foregoing protocol will produce an enhancement inglycemic control and/or insulin sensitivity.

References

[1] Chobanian A V, Bakris G L, Black H R, Cushman W C, Green L A, Izzo JL, Jr. et al. Seventh report of the Joint National Committee onPrevention, Detection, Evaluation, and Treatment of High Blood Pressure.Hypertension 2003; 42(6):1206-1252.

[2] 2003 European Society of Hypertension-European Society of CardiologyGuidelines for the Management of Arterial Hypertension. J Hypertens2003; 21(6):1011-1053.

All patents and publications cited herein are incorporated by referenceinto this application in their entirety.

The words “comprise”, “comprises”, and “comprising” are to beinterpreted inclusively rather than exclusively.

What is claimed is:
 1. A method for delaying progression of diabetic nephropathy in a human subject in need of treatment for diabetic nephropathy comprising: administering to the subject atrasentan or a salt thereof, and a dose of at least one of an angiotensin converting enzyme inhibitor or an angiotensin II receptor blocker, and wherein the method delays the progression of diabetic nephropathy.
 2. The method of claim 1, wherein the method further comprises reducing incidence or severity of adverse cardiac events.
 3. The method of claim 1, comprising administering the dose of the angiotensin converting enzyme inhibitor or angiotensin II receptor blocker at an adequate dose labeled for a hypertension indication.
 4. The method of claim 1, comprising administering the dose of the angiotensin converting enzyme inhibitor or angiotensin II receptor blocker at a highest dose labeled for a hypertension indication.
 5. The method of claim 1, comprising administering the dose of the angiotensin converting enzyme inhibitor or angiotensin II receptor blocker at a highest usual dose prescribed according to Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (JNC 7), British Hypertensive Society (BHD-IV), European Society of Hypertension/European Society of Cardiology (ESH/ESC) or World Health Organization/International Society of Hypertension (WHO/ISH) guidelines.
 6. The method of claim 1, comprising administering the dose of the angiotensin converting enzyme inhibitor or angiotensin II receptor blocker at a highest tolerated dose in the human subject.
 7. The method of claim 1, wherein the angiotensin converting enzyme inhibitor is selected from the group consisting of alacepril, benazepril, captopril, ceronapril, cilazapril, delapril, enalapril, enalaprilat, eosinopril, fosinopril, imidapril, lisinopril, moexipril, moveltipril, omapatrilat, perindopril, quinapril, ramipril, sampatrilat, spirapril, temocapril, trandolapril, and combinations thereof.
 8. The method of claim 1, wherein the angiotensin II receptor blocker is selected from the group consisting of candesartan, eprosartan, irbesartan, losartan, olmesartan, tasosartan, telmisartan, valsartan and combinations thereof.
 9. The method of claim 1, wherein the method further comprises administering a thiazide diuretic selected from the group consisting of althiazide, bendroflumethiazide, benzthiazide, benzylhydrochlorothiazide, buthiazide, chlorothiazide, chlorthalidone, cyclopenthiazide, cyclothiazide, ethiazide, fenquizone, hydrochlorothiazide, hydroflumethiazide, indapamide, methyclothiazide, metolazone, paraflutizide, polythiazide, quinethazone, teclothiazide, trichlormethiazide, and combinations thereof.
 10. The method of claim 1, wherein the method further comprises administering a loop diuretic selected from the group consisting of bumetanide, furosemide, torsemide and combinations thereof.
 11. The method of claim 1, wherein the diabetic nephropathy is overt diabetic nephropathy.
 12. The method of claim 1, comprising administering the atrasentan or a salt thereof for a period of at least about 3 months.
 13. The method of claim 1, comprising administering the atrasentan or a salt thereof for as long as a therapeutic benefit is provided thereby and any adverse side effect thereof remain commensurate with the therapeutic benefit.
 14. The method of claim 1, comprising administering the atrasentan or a salt thereof concomitantly with the angiotensin converting enzyme inhibitor or an angiotensin II receptor blocker.
 15. A method for extending the time to end-stage renal disease or chronic kidney failure in a human subject in need of treatment comprising: administering to the subject atrasentan or a salt thereof, and a dose of at least one of an angiotensin converting enzyme inhibitor or an angiotensin II receptor blocker, and wherein the method extends the time to end-stage renal disease or chronic kidney failure. 